CN104624130A - Method for preparing regenerated chitin microspheres - Google Patents
Method for preparing regenerated chitin microspheres Download PDFInfo
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- CN104624130A CN104624130A CN201510075081.6A CN201510075081A CN104624130A CN 104624130 A CN104624130 A CN 104624130A CN 201510075081 A CN201510075081 A CN 201510075081A CN 104624130 A CN104624130 A CN 104624130A
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- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
- B01J13/02—Making microcapsules or microballoons
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Abstract
The invention relates to a method for preparing regenerated chitin microspheres, belonging to the technical field of high polymer materials. The method comprises the following steps: uniformly mixing purified chitin powder with an alkali aqueous solution, an alkali-urea aqueous solution or an alkali-thiourea aqueous solution, freezing to a freezing point or below, unfreezing at room temperature, repeatedly freezing and unfreezing for 1-2 times, and centrifuging, thereby obtaining a transparent chitin solution, wherein the concentration of chitin is 3wt%-7wt%; dispersing the prepared chitin solution in liquid paraffin containing an emulsifier, wherein the volume of the chitin solution is 10-30 percent of the volume of the liquid paraffin, stirring at the temperature of 0 DEG C until liquid drops are uniformly dispersed, and curing at the temperature of 30-60 DEG C for 5 minutes while stirring; neutralizing by using acid so as to regulate the pH value to be neutral, thereby obtaining the regenerated chitin microspheres; and cleaning the microspheres by using water and ethanol. The nanofiber chitin microspheres have large specific surface area, uniform nanofiber structures and excellent biocompatibility, cells can be subjected to three-dimension adhesion on the surface of the microspheres, and the nanofiber chitin microspheres can be a biological scaffold material with excellent application prospects.
Description
Technical field
The present invention relates to a kind of method preparing regeneration chitin microballoon, belong to technical field of polymer materials.
Background technology
Chitin is that occurring in nature content is only second to cellulosic natural polymer, is extensively present in the cell membrane of shrimp shell, crab shell, insect crust and some fungies, plant.But because chitin is extremely difficult to dissolve, up to the present it remain a kind of and develop minimum biomass resource.And chitin has good biocompatibility and degradable in vivo, it is a kind of very excellent biomaterial.Simultaneously due to the chain rigidity of chitin chain, when the born mutual entanglement again from solution of chitin molecule chain, when forming material, its easy parallel self assembly forms chitin nano fiber, thus prepares chitin nano fiber material.Simultaneously because nanofibrous structures is conducive to the adhesion of cell, and sphere material provides a three-dimensional rack of cell adherence, and it is a kind of cell three-dimensional timbering material having broad prospect of application.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of preparation method regenerating chitin microballoon.The method cost is low, the nanofibers of dimensions narrow distribution of composition chitin microballoon and controlled.
The present invention is using chitin solution as decentralized photo, take isooctane as continuous phase, adopts emulsion method to prepare the narrower regeneration chitin microballoon of Size Distribution.
The present invention by technical solution problem adopts technical scheme is:
(1) be refrigerated to below freezing after the chitin powder of purifying and aqueous alkali, alkali-aqueous solution of urea or alkali-thiourea solution being mixed, at room temperature thaw, repeated freezing-thaw 1-2 time, namely obtains transparent chitin solution after centrifugal, chitin concentration is 3-7wt%;
(2) chitin Solution Dispersion step (1) prepared is in containing in the atoleine of emulsifying agent, the volume of chitin solution is the 10%-30% of atoleine volume, be stirred to after drop is uniformly dispersed at 0 DEG C, solidify 5 minutes at 30-60 DEG C under stirring, then its pH is made to become neutral with acid neutralization, isolate regeneration chitin microballoon, use afterwards water and ethanol purge clean.
As one preferably, chitin solution is dissolved in NaOH-aqueous solution of urea obtained by chitin, and wherein naoh concentration is 2-25wt%, and urea concentration is 1-20wt%, and all the other are water.Preferred naoh concentration is 6-15wt%; Urea concentration is 4-8wt%, and all the other are water.
Described emulsifying agent is one or both the mixture in Tween-85, Span-85.
Narrower its particle diameter of regeneration chitin microballoon of the Size Distribution utilizing said method to prepare is 3 ~ 130 μm, forms the size of its nanofiber between 20-60nm.Prepared nanofiber chitin microballoon can be positioned in 75% alcohol and store, also can stored dry after freeze-drying further.
Patent CN103834069A before inventor is by chitin Solution Dispersion in the paraffin containing emulsifying agent, and solidify at 20 DEG C, add epoxychloropropane, obtain chitin microballoon, size is at more than 200 microns.Because microballoon is larger for this reason, stir easily broken in preparation process, gain in strength as crosslinking agent so we also use epoxychloropropane.And in the present invention, we have changed and have made oil phase with isooctane we can prepare the ball less than CN103834069A size, between several micron to 100 microns, this time, ball would not be broken when stirring.And with high temperature, as 60 degree, can in minutes solidify fast very much.More importantly, ball appearance structure prepared by the ball that the inventive method obtains and CN103834069A is different, the homogeneous structural that the ball in the inventive method is made up of nanofiber, and has the nano-pore of a lot of hundreds of nanometer, and specific area is large, is probably 300m
2/ g, and in CN103834069A, chitin microballoon does not have nanofibrous structures, is made up of the macropore of tens microns, and the hole wall of ball is very smooth, specific area is less only has 20-30m
2/ g.The pattern of chitin microballoon prepared by the inventive method is nanofiber, the nanofibrous microsphere do not reported at present.
The inventive method utilizes alkali/aqueous solution of urea system can realize the dissolving of chitin at low temperatures, simple, nontoxic, the efficient and environmental friendliness of this process operation, and in the dissolving regenerative process of chitin, molecular weight and the acetyl degree of chitin significantly do not change, remain the characteristic of the original excellent biocompatibility of chitin, and homogeneous nanofibrous structures can be prepared by this kind of method, method is very simple, this kind of nanofibrous structures is also extremely conducive to the adhesion of cell, and it can be used as the three-dimensional rack of cell.
Accompanying drawing explanation
Fig. 1 embodiment 3 prepare regeneration chitin microballoon sweep scanning electron microscopic picture (a) and its domain size distribution (a).
Scanning electron microscopic picture on regeneration chitin microballoon prepared by Fig. 2 embodiment 3.
The scanning electron microscopic picture of the three dimensional growth on the regeneration chitin microballoon that Fig. 3 cell is prepared in embodiment 8.
Optical microscope picture on the regeneration chitin microballoon that Fig. 4 cell is prepared in embodiment 8.
Detailed description of the invention
Below will illustrate the present invention by embodiment, but these specific embodiments do not limit the present invention in any way protection domain.Raw material used by the present embodiment all can be buied in market.
Embodiment 1
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 3.3g Tween-85 is added in 500ml there-necked flask, 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 60 DEG C after adding 1.8g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 73 μm of places, its nanofibers of dimensions maximum distribution is at 27nm.Nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 2
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 3.3g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 90ml 7wt% chitin solution after stirring 30min with 500r/min speed and stirs 1 hour, be heated to 60 DEG C after adding 1.8g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 91 μm of places, its nanofibers of dimensions maximum distribution is at 27nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 3
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 6.6.g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 60 DEG C after adding 3.6g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 47 μm of places, its nanofibers of dimensions maximum distribution is at 27nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 4
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 3.3g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 50 DEG C after adding 1.8g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 73 μm of places, its nanofibers of dimensions maximum distribution is at 36nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 5
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 3.3g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 40 DEG C after adding 1.8g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 73 μm of places, its nanofibers of dimensions maximum distribution is at 45nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 6
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 16.5g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 60 DEG C after adding 9g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 20 μm of places, its nanofibers of dimensions maximum distribution is at 27nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 7
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 16.5g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 30ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 60 DEG C after adding 9g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 6 μm of places, its nanofibers of dimensions maximum distribution is at 27nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Embodiment 8
The chitin powder of purifying is added 11wt%NaOH, 4wt% urea and water composition dicyandiamide solution be chilled to-30 DEG C after at room temperature thaw, repeated freezing-course of defrosting 2 times, obtains 7wt% chitin solution.Namely transparent chitin solution is obtained with 6000rpm rotating speed evacuation and centrifugal degassing at 0 DEG C.Under 0 DEG C of ice-water bath, 3.3g Tween-85 is added in 500ml there-necked flask, , 300ml isooctane adds 60ml 7wt% chitin solution after stirring 30min with 1000r/min speed and stirs 1 hour, be heated to 30 DEG C after adding 1.8g Span 85 again and maintain 5min, adding 10% hydrochloric acid again adjusts pH to neutral, sieve chitin balls removing isooctane, repeatedly clean with water and ethanol again, obtain the nanofiber chitin balls that full-size is distributed in 73 μm of places, its nanofibers of dimensions maximum distribution is at 55nm, nanofiber chitin microballoon and liver cell L02 are mixed Dual culture 3 days, cell three dimensional growth thereon can be observed.
Claims (6)
1. regenerate a preparation method for chitin microballoon, it is characterized in that:
(1) be refrigerated to below freezing after the chitin powder of purifying and aqueous alkali, alkali-aqueous solution of urea or alkali-thiourea solution being mixed, at room temperature thaw, repeated freezing-thaw 1-2 time, namely obtains transparent chitin solution after centrifugal, chitin concentration is 3-7wt%;
(2) chitin Solution Dispersion step (1) prepared is in containing in the atoleine of emulsifying agent, the volume of chitin solution is the 10%-30% of atoleine volume, be stirred to after drop is uniformly dispersed at 0 DEG C, solidify 5 minutes at 30-60 DEG C under stirring, then its pH is made to become neutral with acid neutralization, isolate regeneration chitin microballoon, use afterwards water and ethanol purge clean.
2. preparation method according to claim 1, is characterized in that, chitin solution is dissolved in NaOH-aqueous solution of urea obtained by chitin, and wherein naoh concentration is 2-25wt%, and urea concentration is 1-20wt%, and all the other are water.
3. preparation method according to claim 2, is characterized in that, chitin solution is dissolved in NaOH-aqueous solution of urea obtained by chitin, and wherein naoh concentration is 6-15wt%; Urea concentration is 4-8wt%, and all the other are water.
4. preparation method according to claim 1, is characterized in that, described emulsifying agent is one or both the mixture in Tween-85, Span-85.
5. preparation method according to claim 1, is characterized in that, the regeneration chitin microspherulite diameter of preparation is 3 ~ 130 μm, forms the size of its nanofiber between 20-60nm.
6. the chitin microballoon prepared by any one of Claims 1 to 5 is used for the three-dimensional rack of cell.
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Cited By (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN105771823A (en) * | 2016-02-25 | 2016-07-20 | 天津大学 | Method for preparing functional porous micro-spheres |
CN107141494A (en) * | 2017-06-13 | 2017-09-08 | 武汉大学 | A kind of preparation method of chitin nanogel |
CN107158454A (en) * | 2017-05-25 | 2017-09-15 | 福建师范大学 | The preparation method of the porous hemostatic microsphere of chitin |
CN108579630A (en) * | 2018-05-11 | 2018-09-28 | 武汉轻工大学 | The method of pigment in the preparation method and separation grease of chitin nano fiber microballoon |
CN109569590A (en) * | 2018-12-19 | 2019-04-05 | 武汉轻工大学 | The preparation method of chitin base Pd/C catalyst |
CN112973590A (en) * | 2021-03-12 | 2021-06-18 | 四川大学 | Novel preparation method of macroporous chitin microspheres |
CN113336977A (en) * | 2021-05-19 | 2021-09-03 | 武汉大学 | Chitosan nanofiber microsphere and preparation method thereof |
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Publication number | Priority date | Publication date | Assignee | Title |
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CN109569590A (en) * | 2018-12-19 | 2019-04-05 | 武汉轻工大学 | The preparation method of chitin base Pd/C catalyst |
CN112973590A (en) * | 2021-03-12 | 2021-06-18 | 四川大学 | Novel preparation method of macroporous chitin microspheres |
CN113336977A (en) * | 2021-05-19 | 2021-09-03 | 武汉大学 | Chitosan nanofiber microsphere and preparation method thereof |
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