CN106620840A - Silk fibroin modified bone cement porous scaffold and preparation and application thereof - Google Patents
Silk fibroin modified bone cement porous scaffold and preparation and application thereof Download PDFInfo
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- CN106620840A CN106620840A CN201611223915.4A CN201611223915A CN106620840A CN 106620840 A CN106620840 A CN 106620840A CN 201611223915 A CN201611223915 A CN 201611223915A CN 106620840 A CN106620840 A CN 106620840A
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- bone cement
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- fibroin albumen
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/02—Inorganic materials
- A61L27/12—Phosphorus-containing materials, e.g. apatite
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/28—Materials for coating prostheses
- A61L27/34—Macromolecular materials
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/56—Porous materials, e.g. foams or sponges
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2400/00—Materials characterised by their function or physical properties
- A61L2400/18—Modification of implant surfaces in order to improve biocompatibility, cell growth, fixation of biomolecules, e.g. plasma treatment
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/02—Materials or treatment for tissue regeneration for reconstruction of bones; weight-bearing implants
Abstract
The invention relates to a preparation method of a silk fibroin modified bone cement porous scaffold. Raw materials of calcium hydrophosphate and calcium carbonate are fully mixed at the molar ratio of 2:1 by adopting a solid-phase reaction process, the mixture is calcined and rapidly cooled to the room temperature to prepare an alpha-TCP powder, the alpha-TCP powder is taken as a main body, a salt particle with the particle size of 100-300 [mu]m is taken as a pore foaming agent, a disodium hydrogen phosphate solution with the mass fraction of 0.5-5% is taken as a bone cement solidifying solution, the powder is fully mixed, then blended and solidified with the solidifying solution at certain proportion, and soaked in a water bath of 37 DEG C to remove the pore foaming agent, so as to prepare the bone cement porous scaffold; and the porous scaffold is soaked in a silk fibroin solution with the mass fraction of 5-30% for 24 hours, taken out and dried in a drying oven of 60 DEG C to prepare the silk fibroin surface modified bone cement porous scaffold. The silk fibroin modified bone cement porous scaffold meets the requirements of clinical application on the comprehensive performances, has the capability of promoting cell growth, and has a broad application prospect in the fields of clinical bone tissue regeneration.
Description
Technical field
The present invention relates to a kind of method of biology medical material technical field, the modified phosphoric acid of specifically a kind of fibroin albumen
Calcium class bone cement porous support and preparation method and application, are porogen system using salt grain with homemade α-TCP as body of powder
It is immersed in silk fibroin protein solution after standby porous support, drying after taking-up makes rack surface adhere to one layer of fibroin albumen, is prepared into
To the bone cement porous support that fibroin albumen surface is modified.
Background technology
Calcium phosphate bone cement possesses good biocompatibility and osteoinductive due to it, in terms of bone injury reparation
Have broad application prospects.Calcium phosphate bone cement generally by after the powder mixing of two kinds or more synthos with solidify liquid
Lower in certain solid-liquid ratio even, hydration reaction occur, can in room temperature or human body environment self-curing, with free moulding,
The characteristics of solidifying gentle.α-TCP [α phases-tricalcium phosphate] are commonly applied to the powder body of calcium phosphate bone cement, can be with neutral solidification
Liquid room temperature occur hydration reaction, generate main component for hydroxyapatite calcium phosphate bone cement, with human body bone it is inorganic into
Split-phase is seemingly.In the market for the use of calcium phosphate bone cement, a kind of injected bone cement for being to be applied to Minimally Invasive Surgery,
One kind is after external moulding solidification, to prepare the cement brackets such as skeletonization rod, bone piece, and the bone injury for open surgery is repaiied
It is multiple.Wherein, on the one hand calcium phosphate bone cement porous support can provide more due to increased the surface area of material for cell adhesion
Big space, promote bone cell growth, and allow vascular tissue internally grow [Yao Jun, et al,
Biomaterials, 2005], be on the other hand conducive to material to be fully contacted with body fluid, accelerate the degraded of material.
In bone injury repair process, capillary endothelium division, the hypertrophy being wound, to internal extension is damaged, in intravascular
Tube chamber is gradually formed under the impact of blood, then corresponding vascular smooth muscle and adventitia are formed through a series of processes, complete flesh and blood
Pipe regenerative process [Ausprunk DH, MicrovascRec, 1997].Having for reply osteanagenesiss for blood actively promotes
Effect, sufficient blood supply ensure that the release of the blood cell component that fracture repair needs.Fibroin albumen has good biology
The compatibility and biodegradability, the growth to epidermis cell has facilitation, and by support of fibroin albumen material is built
In Tissue Engineering Study, often it is widely used in repairing for the tissues such as blood vessel, skin, artificial ligament as a kind of film healing material
Multiple [Wang Hongxin, Chinese Reconstructive surgery magazine, 2007].
In the research of calcium phosphate bone cement, fibroin albumen is usually used in reinforcing agent and uses, by fibroin fiber and bone water
Compound, the mechanical property of bone cement after increase solidification of mud powder, and advantage of the fibroin albumen in terms of vascular repair is ignored,
Application of the fibroin albumen on Bone Defect Repari is limited significantly.The present invention is directed to background above, is passed using the bone of calcium phosphate bone cement
The property led, the advantage of osteoinductive, in combination with the advantage of fibroin albumen and bone cement loose structure in terms of bone vascular repair, carry
Go out a kind of preparation method of the modified bone cement porous support of fibroin albumen, osteocyte is added while growth is adhered on material
The reparation of fast flesh and blood pipe, and then accelerate the healing of osseous tissue.
The content of the invention
To overcome the deficiencies in the prior art, the present invention to provide a kind of modified bone cement porous support of fibroin albumen and preparation
And application, develop a kind of new bone renovating material for meeting clinical demand.
The purpose of the present invention is achieved through the following technical solutions：It is 2 by mol ratio:1 calcium hydrogen phosphate and Paris white
End is sufficiently mixed；Using solid reaction process, by controlling reaction condition, the α-TCP as bone cement body of powder are prepared
Powder；Grind salt grain and sieve and obtain the salt particle of size 100-300 μm, be sufficiently mixed with α-TCP and prepare bone cement
Powder；Mixing cured with solidify liquid, being put into 37 DEG C of water-bath immersion 24h makes salt grain dissolving obtain bone cement porous support；By this
Frame is put in the silk fibroin protein solution that mass fraction is 5-30% and soaks 24h, is placed in after taking-up in 60 DEG C of baking ovens and is dried, and is prepared into
To the bone cement porous support that fibroin albumen surface is modified.
The preparation method of the modified bone cement porous support of a kind of fibroin albumen, it is characterised in that comprise the steps of：
（1）Using solid reaction process by raw material calcium hydrogen phosphate and Calcium Carbonate in molar ratio 2:After 1 is sufficiently mixed, in 1250-1400
DEG C calcining and is rapidly cooled to room temperature at 2-4h, prepares α-TCP powder, and product Jing wet ball grindings control particle diameter is at 2-4 μm；
（2）It is 100-300 μm of salt grain as porogen with particle diameter based on above-mentioned α-TCP powder, mass fraction is 0.5-5%'s
Disodium phosphate soln reconciles by a certain percentage and solid after powder is sufficiently mixed as bone cement solidify liquid with solidify liquid
Change, the immersion in 37 DEG C of water-baths removes porogen, prepares bone cement porous support；
（3）Above-mentioned porous support is soaked in into 24h in the silk fibroin protein solution that mass fraction is 5-30%, 60 DEG C are placed in after taking-up
It is dried in baking oven, prepares the modified bone cement porous support in fibroin albumen surface.
Step（1）The raw material hybrid mode of described solid reaction process is the wet ball grinding using dehydrated alcohol for medium,
Rotating speed is 400rpm, and Ball-milling Time is 1-4h, and ratio of grinding media to material is mass ratio 3:1.
Step（1）Described product ball milling method is the wet ball grinding using dehydrated alcohol for medium, and rotating speed is 400rpm,
Ball-milling Time is 6h, and ratio of grinding media to material is mass ratio 4:1.
Step（2）Described porogen be it is polished sieve gained, mixed proportion be mass fraction 10-50%, mixing side
Formula is using mortar mixing or dry ball milling.
Step（2）Described bone cement powder and the ratio of solidify liquid is 1.0g/mL-2.0g/mL.
The modified bone cement porous support of a kind of fibroin albumen, it is characterised in that prepared according to any of the above-described methods described
Obtain.
A kind of application of the modified bone cement porous support of fibroin albumen.
The present invention is comprised the following steps：
1st, calcium hydrogen phosphate is mixed homogeneously with Calcium Carbonate, it is mol ratio 2:1.It using dehydrated alcohol is mixed that the hybrid mode is
The ball milling mixing of medium is closed, rotating speed 400rpm, Ball-milling Time is 1-4h, and mixed suspension removes ethanol by rotary evaporation
After be put in 60 DEG C of baking ovens be dried.
2nd, dried calcium hydrogen phosphate and calcium carbonate mixture is calcined in 1250-1400 DEG C of stove and taken out after 2-4h,
Rapidly cool down under air blast environment, obtain α-TCP powder, obtain the α-TCP powder of uniform particle diameter by the way of wet ball grinding afterwards
End, its particle size range is at 2-4 μm.It is standby after drying.
3rd, will sieve after the grinding of crude salt grain, screen out salt particle of the size in 100-300 μ ms.The grinding
Mode can be using mortar grinder or dry ball milling.
4th, disodium phosphate soln of the mass fraction for 0.5-5% is prepared as bone cement solidify liquid.The compound method is
Stirring and dissolving or ultrasonic dissolution assisting under room temperature.
5th, α-TCP powder and salt grain 10-50% in mass ratio is sufficiently mixed, is filled out after mixing by a certain percentage with solidify liquid
Enter mould, 37 DEG C of water-bath immersion 24h are fully put into after solidification, obtain calcium phosphate bone cement porous support.The mixing of the powder
Mode can be using mortar grinder or dry ball milling；The ratio of the powder and solidify liquid is 1.0g/mL-2.0g/mL.
6th, bone cement porous support is put in the silk fibroin protein solution that mass fraction is 5-30% and soaks 24h, taken out rearmounted
It is dried in 60 DEG C of baking ovens, prepares the modified bone cement porous support in fibroin albumen surface.
Using hydroxyapatite bone cement as skeleton, surface is modified using fibroin albumen, with human body bone structure more phase
Seemingly；During Bone Defect Repari, the fibroin albumen surface of material cellular internal is conducive to the adhesion of vascular cell to grow, and accelerates flesh and blood
The reparation of pipe, while osteocyte is also beneficial to be grown in duct, and then promotes the healing of osseous tissue.
The preparation method of the bone cement porous support that the present invention is provided, its is raw materials used simple, and preparation process is simple, can be big
Large-scale production.The modified bone cement porous support of the fibroin albumen meets requirement of the clinical practice to combination property, and possesses rush
Enter the ability of cell growth, have broad application prospects in clinical bone tissue restoration field.
It is an advantage of the current invention that：
1st, the surface for carrying out calcium phosphate bone cement using fibroin albumen is modified so as to the more conducively adhesion of vasculogenic epithelial cells,
With reference to the internal structure of bone cement porous support, accelerate the bone angiogenesiss during bone defect healing, and then promote osseous tissue
Healing.
2nd, preparation method is simple, it is raw materials used simple, it is suitable to a large amount of productions.
Description of the drawings
Fig. 1 is the SEM electron-microscope scanning results of bone cement porous support prepared by this technology, and pore size is 50-300nm.
Fig. 2 is cell proliferation test result of the bone cement porous support prepared by this technology with NIH3T3 cells as object,
Method of testing be embodiment 1 described in, wherein sample 1. -3. respectively correspond to embodiment 2-4 bone cement formula.
Specific embodiment
Following examples are implemented premised on inventive technique scheme, give detailed embodiment and specific behaviour
Make process, but protection scope of the present invention is not limited to following embodiments.
Embodiment 1
（1）It is prepared by α-TCP powder：
In molar ratio 2:1 weighs Dicalcium Phosphate and calcium carbonate powder, the use of appropriate dehydrated alcohol is medium wet ball grinding, turns
Fast 400rpm, Ball-milling Time 4h, ball milling pearl is 3 with powder quality ratio:1.Raw material mixed liquor removes ethanol by rotary evaporation, puts
24h is dried in 60 DEG C of baking ovens.Dried powder is placed in Muffle furnace, is taken out after 1400 DEG C of calcining 2h, in air blast environment
Under rapidly cool down.By the powder after cooling with dehydrated alcohol as medium wet ball grinding, rotating speed 400rpm, Ball-milling Time is 6h, ball
Mill pearl is 4 with powder quality ratio:1.Powder suspension is placed in 80 DEG C of baking ovens and is fully dried, and prepares α-TCP powder.
（2）The preparation of fibroin albumen：
Natural silk degumming is processed, in being dissolved in the mixed solution of calcium chloride, ethanol and deionized water, through dialysis fibroin egg is obtained
White solution, spontaneously dries in 60 DEG C of baking ovens and prepares water miscible silk fibroin powder.
（3）Cell proliferation test method：
Prepare diameter 6mm, the porous support of high 2mm, sterilize 15min in 75% ethanol solution, uses culture medium soaked overnight, examination
Test and 24h in the silk fibroin protein solution of variable concentrations is soaked in after group sample drying, be placed in after taking-up in 60 DEG C of baking ovens and be dried；Control
Group is the blank without sample, as a result such as Fig. 2（0）It is shown.Matched group and test group sample are placed in 96 orifice plates, each
Sample arrange 6 Duplicate Samples, the μ L Deca of cell culture fluid 100 that will be containing 10000/mL of NIH3T3 cells in sample well,
Culture 24h, 48h, 72h.The culture medium incubation 2-4h of the reagents of 10%cck-8 containing volume fraction is added, is measured under 450nm wavelength
Uv absorption, obtains cell proliferation test result.
Embodiment 2
α-TCP prepared by embodiment 1 are 100-300 μm of salt grain in mass ratio 8 with particle diameter：2 mixing, obtain bone cement powder,
It is reconciled with the disodium phosphate soln of mass fraction 2% by the solid-to-liquid ratio of 1.5g/mL, fully 37 DEG C is placed in after solidification
Water-bath immersion 24h removes salt grain, obtains bone cement porous support.Above-mentioned support is soaked in into the fibroin albumen of mass fraction 10%
24h in solution, is placed in 60 DEG C of baking ovens after taking-up and is dried, and obtains the modified bone cement porous support of fibroin albumen.Its SEM Electronic Speculum
Scanning result carries out cell proliferation test, as a result such as Fig. 2 as shown in figure 1, porosity is 38% using the methods described of embodiment 1
（①）It is shown.
Embodiment 3
α-TCP prepared by embodiment 1 are 100-300 μm of salt grain in mass ratio 7 with particle diameter：3 mixing, obtain bone cement powder,
It is reconciled with the disodium phosphate soln of mass fraction 2% by the solid-to-liquid ratio of 1.5g/mL, fully 37 DEG C is placed in after solidification
Water-bath immersion 24h removes salt grain, obtains bone cement porous support.Above-mentioned support is soaked in into the fibroin albumen of mass fraction 10%
24h in solution, is placed in 60 DEG C of baking ovens after taking-up and is dried, and obtains the modified bone cement porous support of fibroin albumen.Porosity is
46%, cell proliferation test is carried out using the methods described of embodiment 1, as a result such as Fig. 2（②）It is shown.
Embodiment 4
α-TCP prepared by embodiment 1 are 100-300 μm of salt grain in mass ratio 7 with particle diameter：3 mixing, obtain bone cement powder,
It is reconciled with the disodium phosphate soln of mass fraction 2% by the solid-to-liquid ratio of 1.5g/mL, fully 37 DEG C is placed in after solidification
Water-bath immersion 24h removes salt grain, obtains bone cement porous support.Above-mentioned support is soaked in into the fibroin albumen of mass fraction 20%
24h in solution, is placed in 60 DEG C of baking ovens after taking-up and is dried, and obtains the modified bone cement porous support of fibroin albumen.Porosity is
45%, cell proliferation test is carried out using the methods described of embodiment 1, as a result such as Fig. 2（③）It is shown.
Claims (7)
1. the preparation method of the modified bone cement porous support of a kind of fibroin albumen, it is characterised in that comprise the steps of：
（1）Using solid reaction process by raw material calcium hydrogen phosphate and Calcium Carbonate in molar ratio 2:After 1 is sufficiently mixed, in 1250-1400
DEG C calcining and is rapidly cooled to room temperature at 2-4h, prepares α-TCP powder, and product Jing wet ball grindings control particle diameter is at 2-4 μm；
（2）It is 100-300 μm of salt grain as porogen with particle diameter based on above-mentioned α-TCP powder, mass fraction is 0.5-5%'s
Disodium phosphate soln reconciles by a certain percentage and solid after powder is sufficiently mixed as bone cement solidify liquid with solidify liquid
Change, the immersion in 37 DEG C of water-baths removes porogen, prepares bone cement porous support；
（3）Above-mentioned porous support is soaked in into 24h in the silk fibroin protein solution that mass fraction is 5-30%, 60 DEG C are placed in after taking-up
It is dried in baking oven, prepares the modified bone cement porous support in fibroin albumen surface.
2. a kind of preparation method of the modified bone cement porous support of fibroin albumen according to claim 1, its feature exists
In step（1）The raw material hybrid mode of described solid reaction process is the wet ball grinding using dehydrated alcohol for medium, and rotating speed is
400rpm, Ball-milling Time is 1-4h, and ratio of grinding media to material is mass ratio 3:1.
3. a kind of preparation method of the modified bone cement porous support of fibroin albumen according to claim 1, its feature exists
In step（1）Described product ball milling method is the wet ball grinding using dehydrated alcohol for medium, and rotating speed is 400rpm, ball milling
Time is 6h, and ratio of grinding media to material is mass ratio 4:1.
4. a kind of preparation method of the modified bone cement porous support of fibroin albumen according to claim 1, its feature exists
In step（2）Described porogen is the polished gained that sieves, and mixed proportion is mass fraction 10-50%, and hybrid mode is
Using mortar mixing or dry ball milling.
5. a kind of preparation method of the modified bone cement porous support of fibroin albumen according to claim 1, its feature exists
In step（2）Described bone cement powder and the ratio of solidify liquid is 1.0g/mL-2.0g/mL.
6. the bone cement porous support that a kind of fibroin albumen is modified, it is characterised in that according to the arbitrary methods described of claim 1-5
Prepare.
7. the application of the modified bone cement porous support of fibroin albumen according to claim 6.
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| CN109678488A (en) * | 2019-01-18 | 2019-04-26 | 华南理工大学 | A kind of ion doping and albumen impregnate dual modified porous calcium phosphate ceramic and preparation method thereof |
| CN114377213A (en) * | 2022-02-25 | 2022-04-22 | 苏州大学附属第一医院 | Novel selenium-enhanced bioactive bone cement and preparation method thereof |
| CN115068686A (en) * | 2022-05-18 | 2022-09-20 | 复旦大学附属中山医院 | Natural-source black phosphorus-silk protein-cellulose biological scaffold |
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| CN107812240B (en) * | 2017-10-19 | 2021-04-09 | 上海纳米技术及应用国家工程研究中心有限公司 | Preparation method of nano-zinc oxide modified antibacterial injectable calcium phosphate bone cement, product and application thereof |
| CN109678488A (en) * | 2019-01-18 | 2019-04-26 | 华南理工大学 | A kind of ion doping and albumen impregnate dual modified porous calcium phosphate ceramic and preparation method thereof |
| CN109678488B (en) * | 2019-01-18 | 2022-01-18 | 华南理工大学 | Ion-doped and protein-impregnated dual-modified porous calcium phosphate ceramic and preparation method thereof |
| CN114377213A (en) * | 2022-02-25 | 2022-04-22 | 苏州大学附属第一医院 | Novel selenium-enhanced bioactive bone cement and preparation method thereof |
| CN115068686A (en) * | 2022-05-18 | 2022-09-20 | 复旦大学附属中山医院 | Natural-source black phosphorus-silk protein-cellulose biological scaffold |
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