CN105561388B - A method of galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds are prepared based on green electrospinning - Google Patents
A method of galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds are prepared based on green electrospinning Download PDFInfo
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- CN105561388B CN105561388B CN201510990389.3A CN201510990389A CN105561388B CN 105561388 B CN105561388 B CN 105561388B CN 201510990389 A CN201510990389 A CN 201510990389A CN 105561388 B CN105561388 B CN 105561388B
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- galactolipin
- chitosan
- native protein
- electrospinning
- nano fiber
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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
- A61L27/14—Macromolecular materials
- A61L27/20—Polysaccharides
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/222—Gelatin
-
- 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
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/227—Other specific proteins or polypeptides not covered by A61L27/222, A61L27/225 or A61L27/24
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
- C08L89/00—Compositions of proteins; Compositions of derivatives thereof
-
- D—TEXTILES; PAPER
- D01—NATURAL OR MAN-MADE THREADS OR FIBRES; SPINNING
- D01D—MECHANICAL METHODS OR APPARATUS IN THE MANUFACTURE OF ARTIFICIAL FILAMENTS, THREADS, FIBRES, BRISTLES OR RIBBONS
- D01D5/00—Formation of filaments, threads, or the like
- D01D5/0007—Electro-spinning
-
- D—TEXTILES; PAPER
- D04—BRAIDING; LACE-MAKING; KNITTING; TRIMMINGS; NON-WOVEN FABRICS
- D04H—MAKING TEXTILE FABRICS, e.g. FROM FIBRES OR FILAMENTARY MATERIAL; FABRICS MADE BY SUCH PROCESSES OR APPARATUS, e.g. FELTS, NON-WOVEN FABRICS; COTTON-WOOL; WADDING ; NON-WOVEN FABRICS FROM STAPLE FIBRES, FILAMENTS OR YARNS, BONDED WITH AT LEAST ONE WEB-LIKE MATERIAL DURING THEIR CONSOLIDATION
- D04H1/00—Non-woven fabrics formed wholly or mainly of staple fibres or like relatively short fibres
- D04H1/70—Non-woven fabrics formed wholly or mainly of staple fibres or like relatively short fibres characterised by the method of forming fleeces or layers, e.g. reorientation of fibres
- D04H1/72—Non-woven fabrics formed wholly or mainly of staple fibres or like relatively short fibres characterised by the method of forming fleeces or layers, e.g. reorientation of fibres the fibres being randomly arranged
- D04H1/728—Non-woven fabrics formed wholly or mainly of staple fibres or like relatively short fibres characterised by the method of forming fleeces or layers, e.g. reorientation of fibres the fibres being randomly arranged by electro-spinning
-
- 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
- A61L2430/28—Materials or treatment for tissue regeneration for liver reconstruction
Abstract
The present invention relates to a kind of methods preparing galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds based on green electrospinning, including：Native protein after purification, obtained aqueous solution are added into galactolipin chitosan, electrospinning obtains galactolipin chitosan-native protein composite nano fiber scaffold.The method of the present invention is simple, and production efficiency is high, has the applications well foreground in terms of bioartificial liver and hepatic tissue reparation；Galactolipin chitosan-native protein/composite nano fiber the biomimetic scaffolds being prepared have good biocompatibility, can promote liver cell adherency and proliferation.
Description
Technical field
The invention belongs to the preparation method field of liver tissue engineering scaffold, more particularly to a kind of green electrospinning prepares half
The method of lactose chitosan-native protein composite nano fiber biomimetic scaffolds.
Background technology
Tissue engineering bracket should simulate the microenvironment of natural extracellular matrix, and the main component of extracellular matrix is glue
Former albumen and polysaccharide are in nanometer fibrous arrangement, and distribution of fiber diameters is in tens to hundreds of nanometers.Therefore, native protein and
The composite nano fiber of polysaccharide composition is believed to preferably simulate natural extracellular matrix.Isinglass, fibroin and chitosan
It is the natural biologic material derived from a wealth of sources, there is excellent biocompatibility, be the preferred material for preparing tissue engineering bracket.It grinds
Study carefully and shows that surface of hepatocytes has a kind of asialoglycoprotein receptor specific recognition galactolipin and can generate specificity therewith mutually
Effect, liver cell show preferably to stick and proliferation behavior on the material containing galactolipin.It will using which kind of preparation method
It is the hot issue in organizational project that holder, which is made, in material, and current existing more mature method includes porous microcarrier, water
Hot method, decellularization and electrostatic spinning etc..And current study show that nano-bracket prepared by electrostatic spinning technique has high hole
Rate, fibre diameter be uniform, mechanical property and biocompatibility have very big advantage, while equipment is simply conducive to safeguard, system
Standby process is seldom or be not related to substantially can be growth factor-loaded to substance, the timbering material that organism and environmental toxic do harm to
The features such as isoreactivity substance, makes electrostatic spinning technique be highly suitable for the research of organizational project and the production of holder.
Invention content
Technical problem to be solved by the invention is to provide one kind preparing galactolipin chitosan-day based on green electrospinning
The method of right albumen composite nano fiber biomimetic scaffolds, this method overcomes the shortcomings of current liver tissue engineering holder, is energy
It is enough more effectively to promote hepatic cell growth and maintain function, the method for the galactolipin chitosan complexes holder of dependable performance.
One kind of the present invention being based on green electrospinning, and to prepare galactolipin chitosan-native protein composite nano fiber bionical
The method of holder, including：
(1) native protein after purification, obtained aqueous solution are added into galactolipin chitosan；Wherein, natural in aqueous solution
The mass concentration of albumen is 10-80wt%, and the mass concentration of galactolipin chitosan is 2-7wt%；
(2) the aqueous solution electrospinning that will be obtained in step (1) obtains galactolipin chitosan-native protein composite nano fiber
Holder；Wherein, electrospinning processes parameter is：Voltage is 9-15KV, fltting speed 0.5-1.5ml/h, and it is 15- to receive distance
25cm, temperature are 20-40 DEG C, humidity 20-30%.
The preparation method of galactolipin chitosan includes in the step (1)：LA is dissolved in the TEMED solution of pH=4.7
(HCl reconciles PH), then activated by the mixture of NHS and EDC；Chitosan is added to the equivalent mole of LA in lysate；Room
Temperature reaction 72h；Product is dialysed, GalCS (galactolipin chitosan) is obtained after freeze-drying.
Native protein is at least one of fibroin and isinglass in the step (1).
The mass concentration of native protein is 10-25wt% in aqueous solution in the step (1).
The post-processing for the galactolipin Properties of Chitosan Fibroin Blend composite nano fiber scaffold that the native protein obtains when being fibroin is adopted
It is stifling with 75% alcohol vapour.
The rear place for galactolipin chitosan-isinglass composite nano fiber scaffold that the native protein obtains when being isinglass
Reason is stifling using 25% glutaraldehyde steam.
Aqueous solution is the aqueous solution that deionized water is prepared in the step (1).
Polyethylene oxide PEO is added in step (1) aqueous solution, the mass concentration of PEO is 0.1wt%-in aqueous solution
0.5wt%..
The present invention prepares galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds using green electrospinning,
The holder has good cell compatibility, can promote adherency and the proliferation of liver cell, be expected in fields such as liver tissue engineerings
Obtain applications well effect.
Advantageous effect
(1) present invention is easy to operate, and experimental provision is simple, and preparation time is short and mild condition, is conducive to produce in enormous quantities；
(2) galactolipin chitosan prepared by the present invention-native protein composite nano fiber biomimetic scaffolds simulation n cell
The structure and composition of epimatrix has good cell compatibility；
(3) galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds prepared by the present invention can quilt due to containing
The gala sugar chain of liver plasma membrane receptor-specific identification is expected to promote adherency and the proliferation of liver cell, is conducive to induction hepatic tissue regeneration
With reconstruction.
Description of the drawings
Fig. 1 is scanning electron microscope (SEM) figure of 1 gained nanofiber biomimetic scaffolds of embodiment；
Fig. 2 is that the SEM of 2 gained nanofiber biomimetic scaffolds of embodiment schemes；
Fig. 3 is that the SEM of liver cell (RH35) growing state on 1 gained nanofiber biomimetic scaffolds of embodiment schemes；
Fig. 4 is that the SEM of liver cell (RH35) growing state on 2 gained nanofiber biomimetic scaffolds of embodiment schemes.
Specific implementation mode
Present invention will be further explained below with reference to specific examples.It should be understood that these embodiments are merely to illustrate the present invention
Rather than it limits the scope of the invention.In addition, it should also be understood that, after reading the content taught by the present invention, people in the art
Member can make various changes or modifications the present invention, and such equivalent forms equally fall within the application the appended claims and limited
Range.
The synthesis step of galactolipin chitosan is in each embodiment：2.3gLA be dissolved in PH=4.7 50mlTEMED/ (by
HCl reconciles PH), then activated by the mixture of NHS (0.14g) and EDC (0.6g)；By 2.2g chitosans (13.7mol) with LA's
Equivalent mole is added in lysate；React at room temperature 72h；Product is dialysed, GalCS is obtained after freeze-drying, and (galactolipin shell is poly-
Sugar).
Embodiment 1
(1) fibroin spinning solution (20wt%) is prepared, mass concentration is added as 0.2% polyethylene oxide (PEO), and according to
Galactolipin chitosan is added in the ratio of mass ratio 1%；
(2) spinning solution in step (1) is transferred in syringe respectively, adjusting voltage is 11KV, and fltting speed is
1.0ml/h, it is 20cm to receive distance, and 30%, temperature is received at 25 DEG C using receiver humidity, and compound fibroin albumen-is made
Galactolipin chitosan nano fiber biomimetic scaffolds.
(3) 75% alcohol vapour of gained nano fibrous membrane is handled can be used afterwards for 24 hours.
Fig. 1 is the SEM figures of the nanofiber biomimetic scaffolds obtained in the present embodiment, and as can be seen from the figure galactolipin shell is poly-
The prepared mixing nano fibrous membrane scaffold fibers surface of sugared (GC) mixing is smooth, and size is uniform, a diameter of 500-750nm
Fig. 3 is the SEM figures of growing state on liver cell (RH35) nanofiber biomimetic scaffolds obtained by the present embodiment, from figure
In it can be seen that liver cell RH35 on GC-SF nano fibrous membranes sticks and to sprawl ground good, it is seen that GC-SF nano fibrous membranes
It is good with the biocompatibility of liver cell.
Embodiment 2
(1) isinglass spinning solution (60wt%) is prepared, galactolipin chitosan is added according to the ratio of mass ratio 3%；
(2) spinning solution in step (1) is transferred in syringe respectively, adjusting voltage is 11KV, and fltting speed is
1.0ml/h, it is 20cm to receive distance, and 30%, temperature is received at 25 DEG C using receiver humidity, and compound isinglass-half is made
Lactose chitosan nano holder.
(3) nanofiber obtained by fumigates 12h with 25% glutaraldehyde steam, is subsequently placed in draught cupboard and divulges information at least
1h is then transferred in vacuum drying chamber dry 2-3 days and can be used.
Fig. 2 is the SEM figures of the nanofiber biomimetic scaffolds obtained in the present embodiment, and what is as can be seen from the figure prepared is compound
Fiber surface is smooth, and diameter is uniform, and the addition of galactolipin-chitosan has no significant effect fiber morphology and diameter, in smooth
Banded structure.
Fig. 4 is the SEM figures of growing state on liver cell (RH35) nanofiber biomimetic scaffolds obtained by the present embodiment, from figure
In it can be seen that liver cell RH35 on GC-Gelatin nano fibrous membranes sticks and sprawls good, it is seen that GC-SF Nanowires
The biocompatibility for tieing up film and liver cell is good.
Embodiment 3
(1) isinglass-fibroin spinning solution (60wt%, 20wt%) is prepared, polyethylene oxide PEO (0.2wt%) is added, and
Galactolipin chitosan is added according to the ratio of mass ratio 1%；
(2) spinning solution in step (1) is transferred in syringe respectively, adjusting voltage is 11KV, and fltting speed is
1.0ml/h, it is 20cm to receive distance, and 30%, temperature is received at 25 DEG C using receiver humidity, and compound isinglass-silk is made
Element-galactolipin chitosan nano holder.
(3) nanofiber obtained by is stifling for 24 hours with 75% alcohol steam, then fumigates 12h with 25% glutaraldehyde steam,
It is subsequently placed in draught cupboard at least 1h that divulges information, is impregnated for several times with deionized water rinse, dry 2-3 is then transferred in vacuum drying chamber
It can be used.
Claims (5)
1. a kind of side preparing galactolipin chitosan-native protein composite nano fiber biomimetic scaffolds based on green electrospinning
Method, including：
(1) native protein after purification, obtained aqueous solution are added into galactolipin chitosan；Wherein, native protein in aqueous solution
Mass concentration be 10-80wt%, the mass concentration of galactolipin chitosan is 2-7wt%；
(2) the aqueous solution electrospinning that will be obtained in step (1) obtains galactolipin chitosan-native protein composite nano fiber scaffold；
Wherein, electrospinning processes parameter is：Voltage is 9-15KV, fltting speed 0.5-1.5ml/h, and it is 15-25cm, temperature to receive distance
It is 20-40 DEG C, humidity 20-30%.
2. it is according to claim 1 it is a kind of prepare galactolipin chitosan-based on green electrospinning native protein be compound receive
The method of rice fiber biomimetic scaffolds, which is characterized in that native protein is at least one in fibroin and isinglass in the step (1)
Kind.
3. it is according to claim 2 it is a kind of prepare galactolipin chitosan-based on green electrospinning native protein be compound receive
The method of rice fiber biomimetic scaffolds, which is characterized in that the native protein is fibroin, and obtained galactolipin Properties of Chitosan Fibroin Blend is multiple
The post-processing for closing nano fiber scaffold is stifling using 75% alcohol vapour.
4. it is according to claim 2 it is a kind of prepare galactolipin chitosan-based on green electrospinning native protein be compound receive
The method of rice fiber biomimetic scaffolds, which is characterized in that the native protein is isinglass, and obtained galactolipin chitosan-fish is bright
The post-processing of glue composite nano fiber scaffold is stifling using 25% glutaraldehyde steam.
5. it is according to claim 1 it is a kind of prepare galactolipin chitosan-based on green electrospinning native protein be compound receive
The method of rice fiber biomimetic scaffolds, which is characterized in that polyethylene oxide PEO is added in step (1) aqueous solution, in aqueous solution
The mass concentration of PEO is 0.1wt% -0.5wt%.
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| CN108676769B (en) * | 2018-04-24 | 2022-09-23 | 武汉仝干医疗科技股份有限公司 | Non-woven fabric support material for promoting growth of liver cells and preparation method thereof |
| CN109529125B (en) * | 2018-12-28 | 2023-01-24 | 佛山科学技术学院 | Method for preparing biological tissue engineering scaffold by solvent spraying |
| CN110882415B (en) * | 2019-11-01 | 2022-02-18 | 中山大学孙逸仙纪念医院 | Composite stent for treating liver injury and preparation method thereof |
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|---|---|---|---|---|
| CN104000739A (en) * | 2014-05-19 | 2014-08-27 | 东华大学 | Preparation method for fish gelatin-silk fibroin composite nanofiber membrane carrying skin-care factor |
| CN104342775A (en) * | 2014-10-15 | 2015-02-11 | 东华大学 | Method for preparing composite nano-fiber membrane with honey and natural materials on basis of environment-friendly electrospinning technology |
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| WO2008118228A2 (en) * | 2006-12-05 | 2008-10-02 | Stonybrook Water Purification | Articles comprising a fibrous support |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104000739A (en) * | 2014-05-19 | 2014-08-27 | 东华大学 | Preparation method for fish gelatin-silk fibroin composite nanofiber membrane carrying skin-care factor |
| CN104342775A (en) * | 2014-10-15 | 2015-02-11 | 东华大学 | Method for preparing composite nano-fiber membrane with honey and natural materials on basis of environment-friendly electrospinning technology |
Non-Patent Citations (1)
| Title |
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| 电纺半乳糖壳聚糖纳米纤维支架及其应用;冯章启 等;《中国化学会第26届学术年会纳米化学分会场论文集》;20080731;106 * |
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