CN106146865A - A kind of dual network high intensity fibroin hydrogel and preparation method thereof - Google Patents
A kind of dual network high intensity fibroin hydrogel and preparation method thereof Download PDFInfo
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
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- A61L2430/40—Preparation and treatment of biological tissue for implantation, e.g. decellularisation, cross-linking
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
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Abstract
The invention belongs to natural macromolecular material technical field, be specially a kind of dual network high intensity fibroin hydrogel and preparation method thereof.The present invention, by will be chemically crosslinked the fibroin hydrogel of (first network) through horseradish peroxidase, is soaked in and can induce fibroinβFold in the solution changed, form physical crosslinking (the second network), the dual network high intensity fibroin hydrogel of solid content 2 20 wt% can be prepared.This hydrogel elastic is good, and intensity is high and is prone to regulation and control, has biocompatibility stable performance under the conditions of simulated in vivo environment, can be widely used for many biomedicines and the regenerative medicine engineering fields such as slow releasing carrier of medication, cartilaginous tissue and ligament tissue.
Description
Technical field
The invention belongs to natural macromolecular material technical field, be specifically related to biological big point of a kind of dual network high-strength natural
Sub-hydrogel and preparation method thereof.
Background technology
Along with the development of field of tissue engineering technology, for excellent performance and be suitable in biological friendly environment the biological doctor used
Grow to even greater heights by the demand of material and technology of preparing thereof.Hydrogel is enriched in the cross-linked network of large quantity of moisture, has stronger leading to
Property thoroughly, and to extracellular matrix the most similar, therefore hydrogel is that the important reconstruction that can be used for of a class is organized
The material of engineering cell support, is again the medicament slow release class carrier material of a great prospect of class.
Fibroin as a kind of natural macromolecular material, its abundance, is provided simultaneously with excellent mechanical property, controlled
Degradation property and good biocompatibility, thus be with a wide range of applications in bio-medical field.But meanwhile,
For the hydrogel material utilizing regenerated silk protein solution to prepare, it is typically due to its mechanical property poor and limit it at some
Actual application in field of tissue engineering technology, such as, need the repair materials such as cartilage or ligament of the bigger pressure of load-bearing.In order to solve this
Problem, studies have reported that and obtain fibroin hydrogel with horseradish peroxidase chemical crosslinking, but its storage modulus generally only has
0.1-10 KPa (CN 104017374 A), and this hydrogel structure is unstable, 37oMeeting is placed for a long time under the conditions of C
The gel transformation to solution occurs.In the research of preparation high intensity fibroin hydrogel, studies have reported that and utilize surface activity
The characteristic of agent prepares the fibroin hydrogel (fibroin/water: 15/85 (w/w)) of high intensity, its modulus of compressibility and stretch modulus
Reach 3.0 MPa, but the fibroin hydrogel prepared when low concentration does not possess good mechanical property, and live in surface
Property agent has certain cytotoxicity;Also studies have reported that and utilize fibroin and cellulose derivative to prepare high-strength composite hydrogel
(CN 104845382 A), the concentration of fibroin is had certain limitations by equally.
The present invention obtains being chemically crosslinked network by horseradish peroxidase chemical crosslinking fibroin, and being then placed on can
Induction fibroin occursβFold and the solution changed is formed physical cross-linked network, obtain that a class modulus is high, toughness strong and has
The well dual network high intensity fibroin hydrogel of biocompatibility, the solid content of this type of hydrogel gel can be at 2-20 wt%
Between adjust control its corresponding mechanical property.In addition, its preparation method is simple, reproducible, and cell can be at it
Surface spreading grows.
Summary of the invention
It is an object of the invention to provide a kind of dual network height with adjustable excellent mechanical performances and good stability
Intensity fibroin hydrogel and preparation method thereof.
The dual network high intensity fibroin hydrogel that the present invention provides, is respectively the fibroin of 2-20 wt% with mass fraction
Aqueous solution forms fibroin hydrogel through horseradish peroxidase chemical crosslinking, is then soaked in and can induce fibroinβFolding
The folded solution changed forms physical crosslinking, obtains dual network high intensity fibroin hydrogel.Described energy inductionβThe water folded
Solution is containing organic solvent, surfactant-based or any one of acid solution.
The preparation method of the dual network high intensity fibroin hydrogel that the present invention provides, concretely comprises the following steps:
(1) silk protein aqueous solution is prepared so that it is ultimate density is 2-20 wt%;
(2) in step (1) gained silk protein solution, horseradish peroxidase (0.01-1 mg/mL) and hydrogen peroxide are added
(0.01-1% v/v) solution, solvent is to be chemically crosslinked at water, 25-50 DEG C, obtains fibroin gel network;
(3) step (2) gained fibroin gel network is soaked in can induce fibroinβFold in the solution changed, during immersion
Between be 6-48 h, obtain dual network high intensity fibroin hydrogel.
In the present invention, described fibroin is Bombyxmori Linnaeus or the one of tussah silk albumen or both mixing.
In the present invention, described horseradish peroxidase is preferably 10:1-1:1 with the quality of hydrogen peroxide than scope.
In the present invention, described can induceβThe solution folded is containing organic solvent, surfactant-based or acid molten
Any one of liquid, or the most several mixing.
In the present invention, described organic solvent class be (but being not limited only to) methanol, ethanol, propanol, butanol, amylalcohol, pyridine,
Any one of acetone, chloroform, or the most several mixing, concentration is 50-95% v/v.
In the present invention, described surfactant-based for (but being not limited only to) sodium lauryl sulphate, DBSA
Any one of sodium or Triton X-100, concentration is 5-40 mM.
In the present invention, described acid solution is any one that pH value of solution can be reduced to 2-5, and concentration is 0.05-5% w/
w。
In the present invention, silk protein aqueous solution described in step (1) can be pure silk protein solution, it is possible to for also containing other compositions
Mixed solution/suspension, other compositions include water soluble polymer, functional inorganic material or drug molecule etc..
The inventive method can also be used for preparing high intensity silk protein membrane and fibroin timbering material.
Dual network high intensity fibroin hydrogel prepared by the present invention, can be used for biomedical engineering and nano-functional material
Field.
Use the inventive method to prepare dual network high intensity fibroin hydrogel, have the advantage that
(1) hydrogel raw material of the present invention is with low cost and through the silk egg of U.S. food Drug Administration certification
In vain, with this gained hydrogel, there is good biocompatibility and biological degradability;
(2) horseradish peroxidase that the present invention uses mainly is extracted from Wasabia japonic (Euterma Wasabi) plant root, realizes commercially producing;Peroxide
Compound is the hydrogen peroxide of low concentration conventional in surgical operation, and has experiment to prove to use the silk of horseradish peroxidase crosslinking
Protein hydrogel no cytotoxicity;
(3) what the present invention used can induce fibroinβFold change solution, and the residual of enzymic catalytic reaction all can by with
Deionized water displacement removes;
(4) the dual network high intensity hydrogel that prepared by the present invention is collectively constituted with physical cross-linked network by being chemically crosslinked network, property
Can be better than by the fibroin hydrogel being individually chemically crosslinked by horseradish peroxidase, and this hydrogel is at simulated in vivo environment bar
Stable performance under part;
(5) present invention still can prepare the hydrogel of mechanics superperformance, difference for the fibroin (such as 2-5 wt%) of low concentration
In the high intensity fibroin hydrogel preparation method the reported restriction to fibroin concentration;
(6) silk protein solution of the present invention, can be water soluble polymer, functional inorganic nanoparticle, drug molecule etc.
Mixed solution, can prepare and there is polyfunctional high intensity hydrogel;
(7) additionally, preparation condition of the present invention is simply efficient, environmental protection, reproducible, there is the value of popularization and application.
Accompanying drawing explanation
Fig. 1 is the Bending Deformation schematic diagram of dual network high intensity fibroin hydrogel (4 wt%).
Fig. 2 is NaHCO3The stretching of fibroin (10 wt%) hydrogel of degumming and the mechanical curves of compression.
Detailed description of the invention
Below in conjunction with specific embodiment, so that the present invention is expanded on further.Be necessary it is pointed out here that, embodiment is served only for
The present invention is further detailed, it is impossible to be interpreted as limiting the scope of the invention, the people that is skilled in technique in this field
Member can make some nonessential improvement and adjustment according to the content of the invention described above.
Embodiment 1
Take Bombyx-mori Silk Proteins solution 2 mL that mass fraction is 10 wt%, add 40 μ L, the horseradish peroxidase of 0.5mg/mL
Enzyme, uniformly adds 40 μ L, 0.05% aqueous hydrogen peroxide solution after mixing, 37 DEG C form gel network;Then 75% it is placed on
Soaking 24 h in ethanol solution, obtain the fibroin hydrogel of high intensity, modulus of compressibility reaches 1.37 MPa, stretch modulus 3.35
MPa.Test shows, stable performance under the conditions of simulated in vivo environment, and cell compatibility is good.
Embodiment 2
Take tussah silk protein solution 4 mL that mass fraction is 2 wt%, add 80 μ L, the horseradish peroxidase of 1 mg/mL,
Uniformly adding 80 μ L, 0.2% aqueous hydrogen peroxide solution after mixing, 40 DEG C form gel network;Then 70% isopropyl it is placed on
Alcohol solution for soaking 12 h, obtains the fibroin hydrogel of high intensity, and modulus of compressibility reaches 0.28 MPa, stretch modulus 0.2
MPa.Test shows, stable performance under the conditions of simulated in vivo environment, and cell compatibility is good.
Embodiment 3
Take tussah silk protein solution 2 mL that mass fraction is 20 wt%, add 40 μ L, the horseradish peroxidase of 0.1 mg/mL
Enzyme, uniformly adds 40 μ L, 0.01% aqueous hydrogen peroxide solution after mixing, 25 DEG C form gel network;Then 0.5M it is placed on
Soaking 48 h in sodium dodecyl sulfate solution, obtain the fibroin hydrogel of excellent mechanical performances, modulus of compressibility reaches 3.2
MPa, stretch modulus 3.5 MPa.Test shows, stable performance under the conditions of simulated in vivo environment, and cell compatibility is good.
Embodiment 4
Take Bombyx-mori Silk Proteins solution 2 mL that mass fraction is 10 wt%, add 40 μ L, the horseradish peroxidase of 0.8 mg/mL
Enzyme, uniformly adds 40 μ L, 0.2% aqueous hydrogen peroxide solution after mixing, 37 DEG C form gel network;Then being placed on pH is
Soaking 24 h in the aqueous acetic acid of 2.5, obtain the fibroin hydrogel of high intensity, modulus of compressibility reaches 1.0 MPa, stretching die
Measure 2.1 MPa.Test shows, stable performance under the conditions of simulated in vivo environment, and cell compatibility is good.
Embodiment 5
Take tussah silk albumen and mixed solution 5 mL of 8 wt% gelatin that mass fraction is 8 wt%, add 100 μ L, 0.5 mg/
The horseradish peroxidase of mL, uniformly adds 100 μ L, 1 % aqueous hydrogen peroxide solution after mixing, 25 DEG C form gel network;
Then it is placed in 0.1M sodium dodecyl sulfate solution and soaks 10 h, obtain the fibroin hydrogel of high intensity, compress mould
Amount reaches 1.25 MPa, stretch modulus 2.0 MPa.Test shows, stable performance under the conditions of simulated in vivo environment, cytocompatibility
Property is good.
Embodiment 6
Take Bombyx-mori Silk Proteins and mixed solution 2 mL of 2 wt% graphene oxides that mass fraction is 12 wt%, add 40 μ L,
The horseradish peroxidase of 0.2 mg/mL, uniformly adds 40 μ L, 0.05% aqueous hydrogen peroxide solution after mixing, 37 DEG C are formed solidifying
Glue network;Then it is placed in 75% butanol solution and soaks 24 h, obtain the functionalization fibroin hydrogel of high intensity, pressure
Contracting modulus reaches 2.52 MPa, stretch modulus 2.35 MPa.Test shows, stable performance under the conditions of simulated in vivo environment, carefully
Born of the same parents' compatibility is good.
Embodiment 7
Take Bombyx-mori Silk Proteins solution 5 mL that mass fraction is 4 wt%, add 50 μ L, the horseradish peroxidase of 1 mg/mL,
Uniformly add 50 μ L, 0.3% aqueous hydrogen peroxide solution after mixing, mixed solution is poured in 10cm*10cm mould, film forming room's warp
Cross 24 hours film forming;Then being placed in 75% ethanol solution and soak 24 h, obtain the silk protein membrane of high intensity, hygrometric state is drop-down
Stretch modulus 103Mpa, elongation strain 300%.Test shows, stable performance under the conditions of simulated in vivo environment, and cell compatibility is good.
Claims (10)
1. the preparation method of a dual network high intensity fibroin hydrogel, it is characterised in that concretely comprise the following steps:
(1) silk protein aqueous solution is prepared so that it is ultimate density is 2-20 wt%;
(2) in step (1) gained silk protein solution, add horseradish peroxidase and the 0.01-1% v/v of 0.01-1 mg/mL
Hydrogenperoxide steam generator, solvent is to be chemically crosslinked at water, 25-50 DEG C, obtains fibroin gel network;
(3) step (2) gained fibroin gel network is soaked in can induce fibroinβFold in the solution changed, during immersion
Between be 6-48 h, obtain dual network high intensity fibroin hydrogel.
Preparation method the most according to claim 1, it is characterised in that described fibroin is Bombyxmori Linnaeus or tussah silk albumen
One or both mixing.
Preparation method the most according to claim 1, it is characterised in that described horseradish peroxidase and hydrogen peroxide
Mass ratio is 10:1-1:1.
4. according to the preparation method described in claim 1,2 or 3, it is characterised in that described can induceβThe solution folded is
Containing any one of organic solvent, surfactant-based or acid solution, or the most several mixing.
Preparation method the most according to claim 4, it is characterised in that described organic solvent class be methanol, ethanol, third
Any one of alcohol, butanol, amylalcohol, pyridine, acetone, chloroform, or the most several mixing, concentration is 50-95% v/v.
Preparation method the most according to claim 4, it is characterised in that described surfactant-based for lauryl sulphate acid
Any one of sodium, dodecylbenzene sodium sulfonate or Triton X-100, concentration is 5-40 mM.
Preparation method the most according to claim 4, it is characterised in that described acid solution is for can be reduced to pH value of solution
Any one of 2-5, concentration is 0.05-5% w/w.
Preparation method the most according to claim 4, it is characterised in that described in step (1), silk protein aqueous solution is pure silk
Protein solution, or be also containing the mixed solution/suspension of other compositions, other compositions include water soluble polymer, functional
Inorganic material or drug molecule.
9. the dual network high intensity fibroin hydrogel that a preparation method as described in one of claim 1-8 prepares.
10. dual network high intensity fibroin hydrogel as claimed in claim 9 is at biomedical engineering and nano-functional material
The application in field.
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Cited By (12)
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CN106420388A (en) * | 2016-11-24 | 2017-02-22 | 江苏爱西施科技服务咨询股份有限公司 | Sunscreen antioxidant silk protein composite hydrogel and preparation method thereof |
CN106673166A (en) * | 2017-01-22 | 2017-05-17 | 厦门大学 | Method for deeply purifying water by using silks |
CN109134889A (en) * | 2018-08-30 | 2019-01-04 | 南通纺织丝绸产业技术研究院 | A kind of photocuring silk fibroin hydrogel and preparation method thereof |
CN109251323A (en) * | 2018-07-25 | 2019-01-22 | 华南理工大学 | Double cross-linked hydrogels of a kind of fibroin albumen-gelatin and preparation method thereof |
CN110144123A (en) * | 2019-04-08 | 2019-08-20 | 复旦大学 | A kind of fibroin/cellulose derivative composite material and preparation method |
CN110227181A (en) * | 2019-05-31 | 2019-09-13 | 武汉大学 | A kind of preparation method and applications of fibroin albumen composite hydroxylapatite material |
CN110229530A (en) * | 2019-07-05 | 2019-09-13 | 青岛大学 | A kind of dual network silk fibroin hydrogel and preparation method thereof |
CN110305339A (en) * | 2019-07-05 | 2019-10-08 | 青岛大学 | A kind of fibroin albumen conductive hydrogel and preparation method thereof |
CN111956871A (en) * | 2020-08-27 | 2020-11-20 | 复旦大学附属中山医院 | Silk protein/gelatin composite material and application thereof |
CN115671388A (en) * | 2022-04-29 | 2023-02-03 | 复向丝泰医疗科技(苏州)有限公司 | Performance-adjustable silk protein injectable microsphere gel and preparation method thereof |
CN115957381A (en) * | 2023-01-30 | 2023-04-14 | 复旦大学附属中山医院 | Injectable sericin hydrogel for regulating bone immune microenvironment |
CN116059450A (en) * | 2023-04-06 | 2023-05-05 | 媄典(北京)医疗器械有限公司 | Preparation method of porous material based on enzyme catalytic crosslinking and porous material |
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WO2015054125A1 (en) * | 2013-10-08 | 2015-04-16 | Trustees Of Tufts College | Tunable covalently crosslinked hydrogels and methods of making the same |
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Patent Citations (1)
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WO2015054125A1 (en) * | 2013-10-08 | 2015-04-16 | Trustees Of Tufts College | Tunable covalently crosslinked hydrogels and methods of making the same |
Cited By (14)
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CN106420388A (en) * | 2016-11-24 | 2017-02-22 | 江苏爱西施科技服务咨询股份有限公司 | Sunscreen antioxidant silk protein composite hydrogel and preparation method thereof |
CN106673166A (en) * | 2017-01-22 | 2017-05-17 | 厦门大学 | Method for deeply purifying water by using silks |
CN109251323A (en) * | 2018-07-25 | 2019-01-22 | 华南理工大学 | Double cross-linked hydrogels of a kind of fibroin albumen-gelatin and preparation method thereof |
CN109134889A (en) * | 2018-08-30 | 2019-01-04 | 南通纺织丝绸产业技术研究院 | A kind of photocuring silk fibroin hydrogel and preparation method thereof |
CN110144123A (en) * | 2019-04-08 | 2019-08-20 | 复旦大学 | A kind of fibroin/cellulose derivative composite material and preparation method |
CN110227181A (en) * | 2019-05-31 | 2019-09-13 | 武汉大学 | A kind of preparation method and applications of fibroin albumen composite hydroxylapatite material |
CN110229530A (en) * | 2019-07-05 | 2019-09-13 | 青岛大学 | A kind of dual network silk fibroin hydrogel and preparation method thereof |
CN110305339A (en) * | 2019-07-05 | 2019-10-08 | 青岛大学 | A kind of fibroin albumen conductive hydrogel and preparation method thereof |
CN110305339B (en) * | 2019-07-05 | 2022-04-26 | 青岛大学 | Silk fibroin conductive hydrogel and preparation method thereof |
CN111956871A (en) * | 2020-08-27 | 2020-11-20 | 复旦大学附属中山医院 | Silk protein/gelatin composite material and application thereof |
CN111956871B (en) * | 2020-08-27 | 2022-06-21 | 复旦大学附属中山医院 | Silk protein/gelatin composite material and application thereof |
CN115671388A (en) * | 2022-04-29 | 2023-02-03 | 复向丝泰医疗科技(苏州)有限公司 | Performance-adjustable silk protein injectable microsphere gel and preparation method thereof |
CN115957381A (en) * | 2023-01-30 | 2023-04-14 | 复旦大学附属中山医院 | Injectable sericin hydrogel for regulating bone immune microenvironment |
CN116059450A (en) * | 2023-04-06 | 2023-05-05 | 媄典(北京)医疗器械有限公司 | Preparation method of porous material based on enzyme catalytic crosslinking and porous material |
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