CN102886070A - Sodium alginate and chitosan skin tissue engineering scaffold material and preparation method thereof - Google Patents
Sodium alginate and chitosan skin tissue engineering scaffold material and preparation method thereof Download PDFInfo
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- CN102886070A CN102886070A CN2012103395390A CN201210339539A CN102886070A CN 102886070 A CN102886070 A CN 102886070A CN 2012103395390 A CN2012103395390 A CN 2012103395390A CN 201210339539 A CN201210339539 A CN 201210339539A CN 102886070 A CN102886070 A CN 102886070A
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Abstract
The invention relates to a sodium alginate and chitosan skin tissue engineering scaffold material and a preparation method thereof. According to the scaffold material, sodium alginate and chitosan take CaCl2 as a crosslinking agent to form a scaffold base, and the mass ratio of the sodium alginate to the chitosan is 5:3-3:5; and a grid structure formed by the sodium alginate is crosslinked by the CaCl2 and put on a sodium alginate and chitosan scaffold base in an interlayer mode, the grid size of the sodium alginate is (1-2)*(1-2)mm<2>, and the height is 0.2 to 0.5mm. By the scaffold material, the sodium alginate and the chitosan base which can be completely degraded are taken as base materials; by combining an interlayer manufactured by a rapid prototyping (RP) technology, the mechanical strength of the sodium alginate and chitosan scaffold material can be improved; and the pore diameter of a scaffold is 100 to 150mu m, the porosity is more than 90 percent, and the tensile strength reaches 1 to 8 MPa. The scaffold material has high biocompatibility and is suitable for growth and proliferation of cells.
Description
Technical field
The present invention relates to a kind of skin tissue engineering bracket material and preparation method thereof, particularly a kind of sodium alginate-chitosan skin tissue engineering bracket material and preparation method thereof.
Background technology
Skin is the organ of human body maximum, is the contacted barrier of human body and external environment, has drainage, feels, prevents moisture evaporation, regulate body temperature, the important functions such as immunity.Skin is self-evident to the importance of human body, yet in actual life, often can be because the reasons such as burn, wound, diabetes chronic ulcer cause skin injury.Any diameter is greater than the full thickness dermal of 4 cm, and human body can't heal fully by self, in these cases, just need to have a kind of suitable wound repair material to promote the healing of wound.Organization engineering skin used 25 years clinically, and for large area skin wounds patient's rehabilitation has brought many benefits, it can fundamentally solve the problem of skin repair, therefore have good development prospect.
At present, skin tissue engineering bracket material mainly is divided into synthesized polymer material and natural macromolecular material.Synthesized polymer material mainly is the polyesters timbering material, and wherein polylactic acid, poly(ethylene oxide) etc. are study hotspots.Although the synthesized polymer material tensile strength is high, degradation rate is controlled because synthetic timbering material lacks the cell recognition signal, be unfavorable for cell adhesion, make its aspect biocompatibility far away not as good as natural macromolecular material.In the natural macromolecular material, collagen gel and collagen sponge are classical artificial skin timbering materials, but their shortcoming is exactly poor mechanical property, and hot strength can only reach 1～4 MPa.
In tissue engineering, sodium alginate and chitosan often are used to prepare tissue engineering bracket material.Lim etc. have prepared a kind of chitosan bilayer skin regeneration support, and by scanning electron microscope and In vitro culture human fibroblast etc. support are estimated.The result shows, the aperture of support is 50-100 μ m, has good cell compatibility, carrying out after cultivating the 14th day of human fibroblast, and cell can be penetrated into internal stent, and quantity begins to increase.
Rapid shaping technique is called for short RP technology (Rapid Prototyping Manufacturing).The RP technology adopts and disperses/the accumulation forming principle, and namely " layering manufacturing, successively stack " is similar to mathematical integral process.Say visually, rapid prototyping system similarly is one " three-dimensional printing machine ".The RP technology is primary for oil-soluble organic polymer rapid shaping at present.
Summary of the invention
One of purpose of the present invention is to provide a kind of sodium alginate-chitosan skin tissue engineering bracket material.This timbering material is biodegradable, has certain mechanical property, toughness and breathability.
Another object of the present invention is to provide the preparation method of this skin tissue engineering bracket material.
A kind of sodium alginate-chitosan skin tissue engineering bracket material, it is characterized in that the structure of this timbering material is: sodium alginate-chitosan is with CaCl
2Form the support matrix as cross-linking agent, wherein the mass ratio of sodium alginate and chitosan is 5:3～3:5; The fenestral fabric that sodium alginate forms is through CaCl
2Crosslinked, place sodium alginate-chitosan support matrix with the interlayer form; Wherein the size of mesh opening of latticed sodium alginate is that 1～2 * 1～2mm, thickness are 0.2～0.5mm.
Above-mentioned timbering material gross thickness be about 0.5～1 mm.
A kind of method for preparing above-mentioned sodium alginate-chitosan skin tissue engineering bracket material is characterized in that the concrete steps of the method are:
A. sodium alginate and chitosan are mixed with the mixed aqueous solution that total solid content is 2～6 wt% by the mass ratio of 5:3～3:5, the pH value of regulating mixed solution is 4～5, obtains the support Matrix Solution;
B. prepare the sodium alginate aqueous solution that the mass percent specific concentration is 2～4 wt%, obtaining size of mesh opening by the RP technology is that 1～2 * 1～2 mm, thickness are the dense grid of 0.2～0.5mm, is 0.5～5% CaCl at mass percent concentration
2Crosslinked 15～45min in the aqueous solution obtains alginic acid grid interlayer;
C. it covers on the alginic acid grid interlayer of step b gained with step a gained support Matrix Solution, leaves standstill 12 hours under-7 ℃ of temperature, moves under 2 ℃ of temperature again and leaves standstill 3～6 hours, under-7 ℃ of temperature, leaves standstill freezing 12 hours at last;
D. the support behind step c gained freezing is put into the freeze dryer lyophilizing, obtain white porous support; Use soak with ethanol, flush away acetic acid, vacuum is drained ethanol; It is immersed in mass percent concentration is 0.5～5% CaCl
2Crosslinked 15～45min in the aqueous solution cleans with deionized water again, namely obtains the sodium alginate-chitosan skin tissue engineering bracket material.
The present invention selects chitosan and sodium alginate as base material.This bi-material is the natural degradable macromolecule, and material source is extensive, and the equal no cytotoxicity of catabolite.Chitosan can be absorbed by the body and not produce toxicity, for the preparation of artificial skin, operation suture thread, artificial kidney film, coagulant, pharmaceutical carrier, and dialyzer, postoperative anti-adhesion membrane and orthopaedics, tooth dental repair material etc.Alginate is a kind of natural polysaccharide that extracts from the Brown algaes such as Thallus Laminariae (Thallus Eckloniae), Macrocystis pyrifera (L.) Ag..It is slightly soluble in water, and safety is nontoxic, and good biocompatibility and immunogenicity are arranged.The sodium alginate-chitosan skin tissue engineering bracket material that adopts the RP technology to prepare has kept natural macromolecular material and has had preferably biocompatibility, the comparatively widely advantage of originating, dense grid interlayer by compound speed forming method preparation makes it have again certain mechanical strength and tear resistance.The present invention prepares water-soluble natural macromolecule mesh-shaped interlayer by rapid shaping technique, is intended to obtain to have the sodium alginate-chitosan skin tissue engineering scaffold of certain mechanical strength and toughness.
Description of drawings
Fig. 1 is the structural representation of gained timbering material of the present invention.
Fig. 2 is the embodiment of the invention three gained 3
#The electron-microscope scanning figure of timbering material.
Fig. 3 fat stem cell is inoculated in the burnt observation figure of the copolymerization after cultivating on the timbering material of the present invention, and wherein a is that fat stem cell is inoculated in 1
#External cultivation burnt observation figure of copolymerization after 3 days on the timbering material.B is that fat stem cell is inoculated in embodiment 1
#External cultivation burnt observation figure of copolymerization after 7 days on the timbering material.C is the b enlarged drawing.Light color is cell among the figure.
The specific embodiment
Embodiment one:
A. preparing total solid content is the support matrix of 4 wt%: the mass ratio of sodium alginate powder and chitosan powder is 5:3, with spirit of vinegar (pH=1～2) regulation system pH value to 4～5.
B. the RP technology prepares interlayer: obtaining size of mesh opening with 2 wt% sodium alginate solns is the dense grid interlayer that 1 * 1 mm, thickness are about 0.5mm, the CaCl 4%
2Crosslinked 30 min in the solution obtain alginic acid grid interlayer.
C. the above-mentioned support Matrix Solution for preparing is poured into and put in the culture dish with dissection, make its grid coverage interlayer.In-7 ℃ of environment, left standstill 12 hours, move into again in 2 ℃ of environment and left standstill 4 hours.At last support is moved on to-7 ℃ of environment, left standstill freezing 12 hours.
D. put into the freeze dryer lyophilizing, take out and use soak with ethanol, vacuum is drained ethanol.CaCl with 4%
2Solution crosslinking 40 min clean with deionized water again, get final product 1
#Timbering material.1
#The average pore size of timbering material is 120 μ m, and porosity is 91.2%, and hot strength is 6.5 MPa.
Embodiment two:
A. preparing total solid content is the support matrix of 4 wt%: the mass ratio of sodium alginate powder and chitosan powder is 5:3, with spirit of vinegar (pH=1～2) regulation system pH value to 4～5.
B. the RP technology prepares interlayer: obtaining size of mesh opening with 4 wt% sodium alginate solns is the dense grid interlayer that 1 * 1 mm, thickness are about 0.5mm, the CaCl 4%
2Crosslinked 30 min in the solution obtain alginic acid grid interlayer.
C. the above-mentioned support Matrix Solution for preparing is poured into and put in the culture dish with dissection, make its grid coverage interlayer.In-7 ℃ of environment, left standstill 12 hours, move into again in 2 ℃ of environment and left standstill 4 hours.At last support is moved on to-7 ℃ of environment, left standstill freezing 12 hours.
D. put into the freeze dryer lyophilizing, take out and use soak with ethanol, vacuum is drained ethanol.CaCl with 4%
2Solution crosslinking 40 min clean with deionized water again, get final product 2
#Timbering material.2
#The average pore size of timbering material is 120 μ m, and porosity is 90.0%, and hot strength is 7.7 MPa.
Embodiment three:
A. preparing total solid content is the support matrix of 4 wt%: the mass ratio of sodium alginate powder and chitosan powder is 5:3, with spirit of vinegar (pH=1～2) regulation system pH value to 4～5.
B. the RP technology prepares interlayer: obtaining size of mesh opening with 4 wt% sodium alginate solns is the dense grid interlayer that 2 * 2 mm, thickness are about 0.5mm, the CaCl 4%
2Crosslinked 30 min in the solution obtain alginic acid grid interlayer.
C. the above-mentioned support Matrix Solution for preparing is poured into and put in the culture dish with dissection, make its grid coverage interlayer.In-7 ℃ of environment, left standstill 12 hours, move into again in 2 ℃ of environment and left standstill 4 hours.At last support is moved on to-7 ℃ of environment, left standstill freezing 12 hours.
D. put into the freeze dryer lyophilizing, take out and use soak with ethanol, vacuum is drained ethanol.CaCl with 4%
2Solution crosslinking 40 min clean with deionized water again, get final product 3
#Timbering material.3
#The average pore size of timbering material is 130 μ m, and porosity is 92.7%, and hot strength is 7.0 MPa.
Embodiment four:
A. preparing total solid content is the support matrix of 4 wt%: the mass ratio of sodium alginate powder and chitosan powder is 1:1, with spirit of vinegar (pH=1～2) regulation system pH value to 4～5.
B. the RP technology prepares interlayer: obtaining size of mesh opening with 4 wt% sodium alginate solns is the dense grid interlayer that 2 * 2 mm, thickness are about 0.5mm, the CaCl 4%
2Crosslinked 30 min in the solution obtain alginic acid grid interlayer.
C. the above-mentioned support Matrix Solution for preparing is poured into and put in the culture dish with dissection, make its grid coverage interlayer.In-7 ℃ of environment, left standstill 12 hours, move into again in 2 ℃ of environment and left standstill 4 hours.At last support is moved on to-7 ℃ of environment, left standstill freezing 12 hours.
D. put into the freeze dryer lyophilizing, take out and use soak with ethanol, vacuum is drained ethanol.CaCl with 4%
2Solution crosslinking 40 min clean with deionized water again, get final product 4
#Timbering material.4
#The average pore size of timbering material is 120 μ m, and porosity is 90.1%, and hot strength is 7.6 MPa.
Illustrate:Fig. 2 is 3
#The Electronic Speculum figure of timbering material, average pore size is about 130 μ m, and the porous perforation, satisfies the cell culture of skin tissue engineering bracket material to the aperture requirement.Fig. 3 a, Fig. 3 b are that fat stem cell is inoculated in 3 days, 7 days figure of external cultivation on the timbering material, and bright green is the cell distribution situation.The timbering material of contrast 3d and 7d can see that the fluorescence scope becomes wide, illustrates that therefore cell breeds at this material.Fig. 3 c is the enlarged drawing of Fig. 3 b, and showed cell is coherent condition at material.
Claims (3)
1. sodium alginate-chitosan skin tissue engineering bracket material, it is characterized in that the structure of this timbering material is: sodium alginate-chitosan is with CaCl
2Form the support matrix as cross-linking agent, wherein the mass ratio of sodium alginate and chitosan is 5:3～3:5; The fenestral fabric that sodium alginate forms is through CaCl
2Crosslinked, place sodium alginate-chitosan support matrix with the interlayer form; Wherein the size of mesh opening of latticed sodium alginate is that 1～2 * 1～2 mm, thickness are 0.2～0.5 mm.
2. sodium alginate-chitosan skin tissue engineering bracket material according to claim 1, it is characterized in that described timbering material gross thickness be about 0.5～1 mm.
3. method for preparing sodium alginate-chitosan skin tissue engineering bracket material according to claim 1 and 2 is characterized in that the concrete steps of the method are:
A. sodium alginate and chitosan are mixed with the mixed aqueous solution that total solid content is 2～6 wt% by the mass ratio of 5:3～3:5, the pH value of regulating mixed solution is 4～5, obtains the support Matrix Solution;
B. prepare the sodium alginate aqueous solution that the mass percent specific concentration is 2～4 wt%, obtaining size of mesh opening by the RP technology is that 1～2 * 1～2 mm, thickness are the dense grid of 0.2～0.5 mm, is 0.5～5% CaCl at mass percent concentration
2Crosslinked 15～45 min in the aqueous solution obtain alginic acid grid interlayer;
C. it covers on the alginic acid grid interlayer of step b gained with step a gained support Matrix Solution, leaves standstill 12 hours under-7 ℃ of temperature, moves under 2 ℃ of temperature again and leaves standstill 3～6 hours, under-7 ℃ of temperature, leaves standstill freezing 12 hours at last;
D. the support behind step c gained freezing is put into the freeze dryer lyophilizing, obtain white porous support; Use soak with ethanol, flush away acetic acid, vacuum is drained ethanol; It is immersed in mass percent concentration is 0.5～5% CaCl
2Crosslinked 15～45 min in the aqueous solution clean with deionized water again, namely obtain the sodium alginate-chitosan skin tissue engineering bracket material.
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Cited By (8)
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| CN103599564A (en) * | 2013-11-21 | 2014-02-26 | 青岛大学 | Preparation method of calcium alginate fibre-reinforced tissue engineering composite scaffold |
| CN104667344A (en) * | 2013-12-03 | 2015-06-03 | 施乐公司 | 3D printing techniques for creating tissue engineering scaffolds |
| CN105148323A (en) * | 2015-08-20 | 2015-12-16 | 东南大学 | Degradable artificial skin scaffold and preparation method thereof |
| CN105169473A (en) * | 2015-10-19 | 2015-12-23 | 东南大学 | Superfine-pore hydrogel bracket and preparation method therefor |
| CN106009943A (en) * | 2016-07-14 | 2016-10-12 | 北京科技大学 | Green, size-controlled, substrate-independent underwater super-oleophobic coating and preparation method thereof |
| CN105999430A (en) * | 2016-07-01 | 2016-10-12 | 赵艳丽 | Composite stent material and preparation method thereof |
| CN107661229A (en) * | 2017-10-26 | 2018-02-06 | 广东海洋大学 | Chitosan and collagen peptide dressing for skin and preparation method thereof |
| CN109847097A (en) * | 2018-12-10 | 2019-06-07 | 安徽徽科生物工程技术有限公司 | Intrinsic bioremediation film, preparation method and application |
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| CN104667344B (en) * | 2013-12-03 | 2020-04-14 | 施乐公司 | 3D printing techniques for producing tissue engineering scaffolds |
| CN105148323A (en) * | 2015-08-20 | 2015-12-16 | 东南大学 | Degradable artificial skin scaffold and preparation method thereof |
| CN105169473A (en) * | 2015-10-19 | 2015-12-23 | 东南大学 | Superfine-pore hydrogel bracket and preparation method therefor |
| CN105999430A (en) * | 2016-07-01 | 2016-10-12 | 赵艳丽 | Composite stent material and preparation method thereof |
| CN106009943A (en) * | 2016-07-14 | 2016-10-12 | 北京科技大学 | Green, size-controlled, substrate-independent underwater super-oleophobic coating and preparation method thereof |
| CN107661229A (en) * | 2017-10-26 | 2018-02-06 | 广东海洋大学 | Chitosan and collagen peptide dressing for skin and preparation method thereof |
| CN109847097A (en) * | 2018-12-10 | 2019-06-07 | 安徽徽科生物工程技术有限公司 | Intrinsic bioremediation film, preparation method and application |
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Application publication date: 20130123 |