CN112843337A - Silk bionic bio-ink and preparation method and application thereof - Google Patents
Silk bionic bio-ink and preparation method and application thereof Download PDFInfo
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- A—HUMAN NECESSITIES
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- 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
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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
- A61L27/14—Macromolecular materials
- A61L27/20—Polysaccharides
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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
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B33—ADDITIVE MANUFACTURING TECHNOLOGY
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- B33Y10/00—Processes of additive manufacturing
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B33—ADDITIVE MANUFACTURING TECHNOLOGY
- B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
- B33Y30/00—Apparatus for additive manufacturing; Details thereof or accessories therefor
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B33—ADDITIVE MANUFACTURING TECHNOLOGY
- B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
- B33Y70/00—Materials specially adapted for additive manufacturing
- B33Y70/10—Composites of different types of material, e.g. mixtures of ceramics and polymers or mixtures of metals and biomaterials
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B33—ADDITIVE MANUFACTURING TECHNOLOGY
- B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
- B33Y80/00—Products made by additive manufacturing
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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
- A61L2400/12—Nanosized materials, e.g. nanofibres, nanoparticles, nanowires, nanotubes; Nanostructured surfaces
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Abstract
The invention discloses a silk bionic bio-ink and a preparation method and application thereof, wherein the silk bionic bio-ink comprises 12-26 wt% of a composite material of silk fibroin and hydroxyapatite, 3-16 wt% of regenerated silk fibroin, 0.5-2 wt% of sodium alginate and 65-75 wt% of water, based on the total weight of the material. The application of the bionic silk ink on 3D printing is printed the shower nozzle and is included outer tube, inner tube and air pressure controlling means, and the inner tube nestification is in the outer tube, and the bionic silk ink is filled to the inner tube, packs the cross-linking curing solution in the outer tube, and air pressure controlling means is used for controlling the flow velocity of the bionic silk ink and the cross-linking curing solution, and the bionic silk ink is extruded together with the cross-linking curing solution in the outer tube, prints out the support in printing the base solution. In the printing process of the support, silk bionic biological ink and a crosslinking curing solution are directly extruded to a printing substrate solution together, crosslinking is carried out to play a role in curing, and steps of heating, spraying an adhesive, ultraviolet curing and the like are not needed.
Description
Technical Field
The invention relates to the technical field of biological scaffold 3D printing, in particular to silk bionic biological ink and a preparation method and application thereof.
Background
3D printing is three-dimensional printing, and compared with traditional machining and other material reduction manufacturing methods, 3D printing is a novel additive manufacturing method. It is a technology for constructing objects by layer-by-layer printing using bondable materials such as powdered metals or plastics based on digital model files. 3D printing is typically achieved using digital technology material printers. The method is often used for manufacturing models in the fields of mold manufacturing, industrial design and the like, and is gradually used for directly manufacturing some products, and parts printed by the technology are already available. The technology has applications in jewelry, footwear, industrial design, construction, engineering and construction, automotive, aerospace, dental and medical industries, education, geographic information systems, civil engineering, firearms, and other fields. In addition, in recent years, 3D printing technology has been widely used for biomedical materials.
At present, the forming method of the 3D printer to the material in the printing process mainly includes several methods, such as laser light solidification, fused deposition forming, adhesive spraying, laser sintering, and the like. During the forming process, additional equipment or adhesive material needs to be introduced.
Disclosure of Invention
The invention aims to overcome the defects of the prior art, and provides the silk bionic bio-ink which is matched with a crosslinking curing solution for 3D printing, directly plays a role in curing a printing support, does not need steps of heating, spraying an adhesive, ultraviolet curing and the like, and has the advantages of simplicity and easiness in operation.
The invention also aims to provide a preparation method of the silk bionic bio-ink.
The invention further aims to provide application of the silk bionic bio-ink in 3D printing.
The technical scheme of the invention is as follows: the silk bionic bio-ink comprises, by taking the total weight of the material as a reference, 12-32.5 wt% of a composite material of silk fibroin and hydroxyapatite, 2-16 wt% of regenerated silk fibroin, 0.5-2 wt% of sodium alginate and 65-75 wt% of water.
The other technical scheme of the invention is as follows: a preparation method of silk bionic bio-ink comprises the steps of putting 12-32.5 parts by weight of a silk fibroin and hydroxyapatite composite material, 0.5-2 parts by weight of sodium alginate, 2-16 parts by weight of regenerated silk fibroin and 65-75 parts by weight of water into a beaker for mixing, and stirring at room temperature for 12-14 hours.
Further, the preparation of the silk fibroin and hydroxyapatite composite material comprises the following steps:
step S1: putting the silk fibroin obtained after degumming into a prepared ternary system, and heating and dissolving to obtain a silk fibroin solution;
step S2: adding diammonium hydrogen phosphate aqueous solution into the silk fibroin solution obtained in the step S1, adjusting the reaction system to be alkaline by ammonia water, and reacting at the temperature of 60-80 ℃ for 12-24 hours;
step S3: and (4) after the reaction product in the step (S2) is filtered and washed, freeze-drying the reaction product by using a freeze dryer to obtain a powdery silk fibroin and hydroxyapatite composite material, wherein hydroxyapatite nanorods in the composite material grow on a silk fibroin substrate.
Further, in step S1, the degumming step is to heat a sodium carbonate solution with a mass fraction of 0.5% to a slight boiling state, and then add silk, the mass M of the silk: stirring the solution with a glass rod for 0.5h when the volume V of the solution is 1:100, washing the degummed solution with deionized water, repeating the above work to carry out secondary degummed treatment, and airing the degummed solution at room temperature.
Further, the ternary system is prepared from calcium chloride, ethanol and water, wherein the molar ratio of the calcium chloride to the ethanol to the water is 1 (1.8-2.2) to (7.5-8.5).
Further, in step S2, the amount of diammonium phosphate is such that the molar ratio of Ca to P is (1.65-1.69):1, and the PH of the system is 10-12 after adjustment with ammonia, based on the Ca element contained in the silk fibroin solution and the P element in the diammonium phosphate aqueous solution.
The invention also adopts the technical scheme that: an application of the silk bionic bio-ink in 3D printing.
Further, including printing the shower nozzle, print the shower nozzle and include outer tube, inner tube and air pressure controlling means, the inner tube nestification is in the outer tube, and the bionical biological ink of silk is filled in the inner tube, fills the cross-linking curing solution in the outer tube, and air pressure controlling means is used for controlling the flow velocity of the bionical biological ink of silk and cross-linking curing solution, and the bionical biological ink of silk in the inner tube and the cross-linking curing solution in the outer tube are extruded together, prints out the support in printing the base solution.
Further, the crosslinking curing solution contains ethanol and calcium chloride solution in a volume ratio of 1:3 based on the volume ratio.
Further, the preparation method of the printing substrate solution comprises the following steps: 0.004g of horseradish peroxidase was added to 150ml of water, dissolved with stirring, and finally 50ml of ethanol and 1.5ml of hydrogen peroxide were added.
Compared with the prior art, the invention has the following beneficial effects:
according to the silk bionic bio-ink disclosed by the invention, in the printing process of the stent, the silk bionic bio-ink and the crosslinking curing solution are directly extruded to the substrate solution together, and crosslinking is carried out to play a role in curing, so that the steps of heating, spraying an adhesive, ultraviolet curing and the like are not required.
According to the application of the silk bionic bio-ink in 3D printing, the supports with different shapes are obtained by printing the blended multiple materials layer by using a 3D direct-writing printing technology, the direct-writing printing has the advantages of short time consumption, low cost, controllable pore size, shape and connectivity of the supports, low requirements on temperature and environment, simplicity in operation and the like.
Drawings
Fig. 1 is a scanning electron microscope image of the silk fibroin and hydroxyapatite composite material of the present invention.
Fig. 2 is a schematic structural diagram of a print head according to the present invention.
Fig. 3 is a schematic view of the cross-linking effect of the silk bionic bio-ink and the cross-linking curing solution of the invention.
Fig. 4 shows the scaffolds obtained by printing in examples 2, 3 and 4 of the present invention, which are a grid-shaped scaffold, a hollow cylindrical scaffold and a honeycomb-shaped scaffold from left to right.
An outer tube 1, an inner tube 2, a printing substrate solution 3.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto.
Example 1
The embodiment provides silk bionic bio-ink, which contains 12-32.5 wt% of a composite material of silk fibroin and hydroxyapatite, 2-16 wt% of regenerated silk fibroin, 0.5-2 wt% of sodium alginate and 65-75 wt% of water, based on the total weight of the material.
In this embodiment, the preparation method of the silk bionic bio-ink comprises: weighing 0.5g of regenerated silk fibroin, placing the regenerated silk fibroin in a beaker, adding 16g of water for dissolving, then adding 0.4g of sodium alginate, after the sodium alginate is dissolved, adding 5g of a silk fibroin and hydroxyapatite composite material, stirring the mixture at room temperature by using a magnetic stirrer for 12 hours, and fully dispersing the hydroxyapatite in a mixed solution of the regenerated silk fibroin and the sodium alginate to obtain the silk bionic bio-ink.
As shown in fig. 1, the preparation of the composite material of silk fibroin and hydroxyapatite comprises the following steps:
step S1: degumming silk fibroin, heating a sodium carbonate solution with the mass fraction of 0.5% to slight boiling, and then adding silk, wherein the mass M of the silk is as follows: stirring the solution with a glass rod for 0.5h with the volume V of 1:100, washing with deionized water after degumming, repeating the above steps to carry out secondary degumming treatment, and airing at room temperature after degumming; putting the silk fibroin obtained after degumming into a prepared ternary system, wherein the ternary system is prepared from calcium chloride, ethanol and water, the molar ratio of the calcium chloride to the ethanol to the water is 1:2:8, and heating and dissolving to obtain a silk fibroin solution;
step S2: adding diammonium hydrogen phosphate aqueous solution into the silk fibroin solution obtained in the step S1, adjusting the reaction system to be alkaline by ammonia water, and reacting at the temperature of 60-80 ℃ for 12-24 hours;
step S3: and (4) after the reaction product in the step (S2) is filtered and washed, freeze-drying the reaction product by using a freeze dryer to obtain a powdery silk fibroin and hydroxyapatite composite material, wherein hydroxyapatite nanorods in the composite material grow on a silk fibroin substrate.
Example 2
As shown in fig. 2 and 3, this embodiment is the application of the bionic biological ink of silk in embodiment 1 on 3D printing, including printing the shower nozzle, it includes outer tube 1, inner tube 2 and air pressure controlling means to print the shower nozzle, and the inner tube nestification is in the outer tube, packs the bionic biological ink of silk in the inner tube, packs cross-linking curing solution in the outer tube, and air pressure controlling means is used for controlling the flow velocity of the bionic biological ink of silk and cross-linking curing solution, and the bionic biological ink of silk in the inner tube and the cross-linking curing solution in the outer tube are extruded together, prints out the support in printing base solution 3.
The crosslinking curing solution contains ethanol and calcium chloride solution in a volume ratio of 1:3 based on the volume ratio. During preparation, 10g of calcium chloride and 150ml of water are added into a beaker and stirred and dissolved by using a magnetic stirrer to obtain a calcium chloride solution. Adding 50ml of ethanol and 150ml of calcium chloride solution into another beaker, and uniformly mixing to obtain a crosslinking curing solution.
The preparation method of the printing substrate solution comprises the following steps: 0.004g of horseradish peroxidase was added to 150ml of water, dissolved with stirring, and finally 50ml of ethanol and 1.5ml of hydrogen peroxide were added.
As shown in fig. 4, during printing, a latticed scaffold program is written on a direct-writing printing device, the silk bionic biological ink prepared in example 1 is filled into an inner tube, air is centrifugally exhausted, a cross-linking curing solution is added into an outer tube, an air pressure control device is adjusted to enable the pressure to be 15Psi and the linearity to be 8mm/s, the silk bionic biological ink in the inner tube and the cross-linking curing solution in the outer tube are extruded into a printing base solution together, and 3D layer-by-layer printing is carried out to manufacture a latticed scaffold. The obtained scaffold was subjected to a compression test by an electronic universal material testing machine, and the compressive strength was 60 MPa.
Example 3
The difference between this example and example 2 is that, as shown in fig. 4, during printing, a hollow cylinder program is written on a direct-writing printing device, the silk bionic bio-ink prepared in example 1 is filled into an inner tube, air is centrifugally exhausted, the cross-linking curing solution in example 2 is added into an outer tube, an air pressure control device is adjusted so that the pressure is 15Psi and the linear degree is 8mm/s, the silk bionic bio-ink in the inner tube and the cross-linking curing solution in the outer tube are extruded into a printing base solution together, and 3D layer-by-layer printing is performed to prepare the hollow cylinder stent. The obtained support is subjected to a compression test by an electronic universal material testing machine, and the measured compression strength is 65 MPa.
Example 4
The difference between this example and example 2 is that, as shown in fig. 4, during printing, a honeycomb scaffold program is written on a direct-writing printing device, the silk bionic bio-ink prepared in example 1 is filled into an inner tube, air is centrifugally exhausted, the cross-linking curing solution in example 2 is added into an outer tube, an air pressure control device is adjusted so that the pressure is 15Psi and the linear degree is 8mm/s, the silk bionic bio-ink in the inner tube and the cross-linking curing solution in the outer tube are extruded into a printing base solution together, and 3D layer-by-layer printing is performed to prepare the honeycomb scaffold. The obtained support is subjected to a compression test by an electronic universal material testing machine, and the measured compression strength is 70 MPa. In other embodiments, the stent may also be a strip or other shape, all falling within the scope of the present application and not further listed here.
Example 5
The difference between the embodiment and the embodiment 1 is that the preparation method of the silk bionic bio-ink comprises the following steps: weighing 2g of regenerated silk fibroin, placing the regenerated silk fibroin in a beaker, adding 16g of water for dissolving, then adding 0.4g of sodium alginate, after the sodium alginate is dissolved, adding 5g of a composite material of silk fibroin and hydroxyapatite, stirring the mixture at room temperature by using a magnetic stirrer for 13 hours, and fully dispersing the hydroxyapatite in a mixed solution of the regenerated silk fibroin and the sodium alginate to obtain the silk bionic bio-ink.
Example 6
The difference between the embodiment and the embodiment 1 is that the preparation method of the silk bionic bio-ink comprises the following steps: the preparation method of the silk ink comprises the following steps: weighing 0.5g of regenerated silk fibroin, placing the regenerated silk fibroin in a beaker, adding 16g of water for dissolving, then adding 0.1g of sodium alginate, after the sodium alginate is dissolved, adding 5g of a silk fibroin and hydroxyapatite composite material, stirring the mixture at room temperature by using a magnetic stirrer for 14 hours, and fully dispersing the hydroxyapatite in a mixed solution of the regenerated silk fibroin and the sodium alginate to obtain the silk bionic bio-ink.
As mentioned above, the present invention can be better realized, and the above embodiments are only preferred embodiments of the present invention, and are not intended to limit the scope of the present invention; all equivalent changes and modifications made according to the present disclosure are intended to be covered by the scope of the claims of the present invention.
Claims (10)
1. The silk bionic bio-ink is characterized by comprising 12-32.5 wt% of a composite material of silk fibroin and hydroxyapatite, 2-16 wt% of regenerated silk fibroin, 0.5-2 wt% of sodium alginate and 65-75 wt% of water, wherein the total weight of the material is taken as a reference.
2. A preparation method of silk bionic bio-ink is characterized in that 12-32.5 parts by weight of composite material of silk fibroin and hydroxyapatite, 0.5-2 parts by weight of sodium alginate, 2-16 parts by weight of regenerated silk fibroin and 65-75 parts by weight of water are placed into a beaker for mixing, and stirring is carried out at room temperature for 12-14 hours.
3. The method for preparing silk biomimetic bio-ink according to claim 2, wherein the preparation of the silk fibroin and hydroxyapatite composite material comprises the following steps:
step S1: putting the silk fibroin obtained after degumming into a prepared ternary system, and heating and dissolving to obtain a silk fibroin solution;
step S2: adding diammonium hydrogen phosphate aqueous solution into the silk fibroin solution obtained in the step S1, adjusting the reaction system to be alkaline by ammonia water, and reacting at the temperature of 60-80 ℃ for 12-24 hours;
step S3: and (4) after the reaction product in the step (S2) is filtered and washed, freeze-drying the reaction product by using a freeze dryer to obtain a powdery silk fibroin and hydroxyapatite composite material, wherein hydroxyapatite nanorods in the composite material grow on a silk fibroin substrate.
4. The method for preparing silk biomimetic bio-ink according to claim 3, wherein in the step S1, the degumming step is that a sodium carbonate solution with a mass fraction of 0.5% is heated to a slight boiling state, and then silk, the mass M of which is: stirring the solution with a glass rod for 0.5h when the volume V of the solution is 1:100, washing the degummed solution with deionized water, repeating the above work to carry out secondary degummed treatment, and airing the degummed solution at room temperature.
5. The preparation method of the silk biomimetic bio-ink according to claim 3, characterized in that the ternary system is prepared from calcium chloride, ethanol and water, and the molar ratio of calcium chloride, ethanol and water is 1 (1.8-2.2) to (7.5-8.5).
6. The method for preparing silk bionic bio-ink according to claim 3, wherein in step S2, the amount of diammonium phosphate is such that the molar ratio of Ca/P is (1.65-1.69):1, and the pH of the system is 10-12 after ammonia adjustment, calculated as Ca element contained in the silk fibroin solution and P element in the diammonium phosphate aqueous solution.
7. Use of the silk biomimetic bio-ink of claim 1 in 3D printing.
8. The application of the silk bionic biological ink in 3D printing according to claim 1, which comprises a printing nozzle, wherein the printing nozzle comprises an outer tube, an inner tube and an air pressure control device, the inner tube is nested in the outer tube, the silk bionic biological ink is filled in the inner tube, the cross-linking curing solution is filled in the outer tube, the air pressure control device is used for controlling the outflow speed of the silk bionic biological ink and the cross-linking curing solution, the silk bionic biological ink in the inner tube and the cross-linking curing solution in the outer tube are extruded together, and a support is printed in a printing substrate solution.
9. The application of the silk biomimetic bio-ink to 3D printing according to claim 8, wherein the crosslinking and curing solution contains ethanol and calcium chloride in a volume ratio of 1:3 based on the volume ratio.
10. The application of the silk biomimetic bio-ink in 3D printing according to claim 8, wherein the preparation method of the printing substrate solution comprises the following steps: 0.004g of horseradish peroxidase was added to 150ml of water, dissolved with stirring, and finally 50ml of ethanol and 1.5ml of hydrogen peroxide were added.
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Cited By (1)
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| WO2023152202A1 (en) * | 2022-02-10 | 2023-08-17 | Amsilk Gmbh | Gradient printing reservoir and printing method |
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| US20190160203A1 (en) * | 2016-06-03 | 2019-05-30 | Cellink Ab | Preparation and applications of rgd conjugated polysaccharide bioinks with or without fibrin for 3d bioprinting of human skin with novel printing head for use as model for testing cosmetics and for transplantation |
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| WO2020223415A1 (en) * | 2019-04-29 | 2020-11-05 | Trustees Of Tufts College | Systems and methods for living silk articles |
| CN110302419A (en) * | 2019-06-19 | 2019-10-08 | 暨南大学 | A kind of 3D printing compound bio ink material and the preparation method and application thereof |
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| WO2023152202A1 (en) * | 2022-02-10 | 2023-08-17 | Amsilk Gmbh | Gradient printing reservoir and printing method |
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