CN1448425A - Process of preparing polymer porous leg by separation and combination of paraffin microballons porous making agent and thermotropic phase - Google Patents
Process of preparing polymer porous leg by separation and combination of paraffin microballons porous making agent and thermotropic phase Download PDFInfo
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- CN1448425A CN1448425A CN 03116446 CN03116446A CN1448425A CN 1448425 A CN1448425 A CN 1448425A CN 03116446 CN03116446 CN 03116446 CN 03116446 A CN03116446 A CN 03116446A CN 1448425 A CN1448425 A CN 1448425A
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Abstract
The production process of porous polymer rack includes suspending dispersion to prepare paraffin microballoon, mixing paraffin microballoon and polymer solution, lowering temperature to make polymer solution produce phase separation, and freeze drying to eliminate solvent, boiled hexane washing or extracting to eliminate paraffin microballoon. The said process concentrates the advantages of both pore creating process and thermal phase separation process and the produced porous polymer rack has macro pores with size determined by the paraffin microballoon as pore creating agent and micro pores produced in thermal phase separation and distributed on wall of the macro pores. The porous polymer rack has controllable pore size, good communication and high porosity.
Description
Technical field
The present invention relates to the preparation method of used in tissue engineering porous polymer scaffold, relate to wax ball pore-foaming agent specifically and be separated to combine with thermic and prepare the method for used in tissue engineering porous polymer scaffold.
Background technology
Damaged or the nonfunction of tissue is common disease, has had a strong impact on patient's healthy and life quality.To one of modern treatment method of these diseases is tissue engineering technique.The main thought of organizational project is inoculated on the extracellular matrix surrogate for the viable cell that will have certain correlation function, and this surrogate can be cell a space structure is provided, and cell is grown in the above.Vitro culture forms the mixture of cell and surrogate after the certain period, the gained mixture is transplanted to damaged tissue place in the body again, to repair damaged tissue.Used alternative materials is generally biodegradable artificial-synthetic copolymer's timbering material or natural biomacromolecule timbering material in the organizational project.
Because have that degradation speed is controlled, advantages such as good mechanical performance and convenient sources, biodegradable polymer porous support material has obtained widespread use in organizational project.Organizational project for the requirement of porous polymer scaffold microtexture is: have suitable aperture, higher porosity and good hole connectedness, so that the entering of cell, the infiltration of nutritive substance and the discharge of refuse.
The preparation of used in tissue engineering porous polymer scaffold at present has several different methods, as non-braiding shape fiber-forming method, pore-creating agent method, phase separation method, foaming and three dimensional printing method etc.Wherein pore-creating agent method and thermally induced phase separation all have easy to operately, do not need the advantage of specific installation.The pore-creating agent ratio juris is: pore-creating agent is mixed with polymers soln, pore-creating agent is leached the formation porous support materials behind the removal solvent again.The principle of thermally induced phase separation is: polymers soln is cooled to it below phase-splitting point, polymers soln is divided into is rich in polymkeric substance and the two-phase that is rich in solvent, quenching is fixed the form of phase region then, at last solvent phase is removed, and obtains porous support materials.For the pore-creating agent method, its advantage is that the aperture of porous support can be controlled by the particle diameter of control pore-creating agent.Yet utilize the pore-creating agent method merely,, exist the tediously long shortcoming of preparation process, and porosity is lower in order to obtain having the support of good hole connectedness.For thermally induced phase separation, its advantage is that the porous support that is obtained has good hole connectedness, is difficult for accurately control yet have the aperture, the especially difficult shortcoming that obtains the large size aperture.
Summary of the invention
The present invention seeks to for obtain having suitable aperture, the used in tissue engineering porous polymer scaffold of height porosity and good hole connectedness, proposed paraffin microballons porous making agent method and thermally induced phase separation and combined and prepare the method for used in tissue engineering porous polymer scaffold.
Method of the present invention specifically may further comprise the steps:
(1) adopt the suspended dispersed legal system to be equipped with the paraffin microballoon that particle diameter is 50 μ m～700 μ m;
(2) polymers soln that the paraffin microballoon that obtains and concentration are 0.01-0.5g/mL is under 15～55 ℃ of temperature, mixed by 0.5～3g paraffin microballoon/mL polymers soln is even, reduce temperature then to-100～10 ℃, polymers soln is separated;
(3) solvent in the polymers soln is removed in lyophilize, obtains paraffin microballoon and mixture of polymers;
(4) above-mentioned paraffin microballoon and mixture of polymers are placed the washing of ebullient normal hexane repeatedly, perhaps utilize cable-styled extractor that the extracting of paraffin microballoon is removed;
(5) normal hexane is removed in vacuum-drying, obtains porous polymer scaffold.
Among the present invention, suspension method prepares the paraffin microballoon and can adopt: with solid paraffin and concentration is the aqueous dispersant mixing of 1～100mg/mL, solid paraffin/aqueous dispersant ratio is 1～100g/100mL, be heated to 60 ℃～100 ℃, make paraffin become liquid state, stir, the paraffin of liquid state is separated into small droplets, after treating paraffin suspension in stable condition, pour quenching in-5 ℃～10 ℃ the cold water fast into, obtain solid paraffin microballoon.
The dispersion agent that above-mentioned suspension method prepares the paraffin microballoon comprises the tensio-active agent that gelatin, polyvinyl alcohol, polyoxyethylene glycol, sodium laurylsulfonate etc. are commonly used.According to the concentration of used surfactant soln and the difference of paraffin consumption, prepared wax spherolite directly has different distributions.Adopt the domestic standard sieve that the paraffin microballoon is carried out classification, with the paraffin microballoon by size be divided into below the 150 μ m, not at the same level part of 150～280 μ m, 280～450 μ m and 450～700 μ m etc.The consumption of the particle diameter by control paraffin microballoon, the concentration of polymers soln and paraffin microballoon can obtain the porous polymer scaffold of different apertures and porosity.
Among the present invention, said polymkeric substance mainly is but is not limited to poly(lactic acid) (PLA), comprise polyglycolic acid (PGA), lactic acid-ethanol copolymer (PLGA), polycaprolactone (PCL), urethane (PU), polycarbonate, poly-acid anhydrides, poly phosphazene, poly butyric ester and multipolymer thereof, collagen, chitosan etc. and above-mentioned mixture of polymers and the multipolymer between them, the product that also comprises the different spaces structure of above-mentioned polymkeric substance, as Poly-L-lactic acid (PLLA), dextrorotation poly(lactic acid) (PDLA) and meso poly(lactic acid) (PD, LLA).
In the step of the present invention (2), phase separation temperature is according to different polymers solns and difference.For the dioxane solution of poly--L-lactic acid, phase separation temperature is between 10～-20 ℃.
The present invention has concentrated the advantage of two kinds of methods by wax ball method is combined with thermally induced phase separation, has obtained having suitable aperture, the used in tissue engineering porous polymer timbering material of height porosity and good hole connectedness.A notable feature of the porous polymer scaffold that the present invention is prepared is the size decision of big hole dimension by pore-creating agent paraffin microballoon, the aperture that the thermic that also distributing in the hole wall between macropore is separated and is produced, so porosity height.That the present invention has is easy to operate, controllable aperture, the hole connectedness is good, porosity is high advantage.
Description of drawings
Fig. 1 utilizes combine with the thermally induced phase separation stereoscan photograph of PLLA porous support of preparation of paraffin microballons porous making agent method.Wax spherolite footpath 150～450 μ m, PLLA concentration 0.06g/mL, wax ball consumption 1.3g/mL polymers soln.
Fig. 2 a is the optical microscope photograph of diameter less than the paraffin microballoon of 150 μ m.
Fig. 2 b is that diameter is the optical microscope photograph of the paraffin microballoon of 150～280 μ m.
Fig. 2 c is that diameter is the optical microscope photograph of the paraffin microballoon of 280～450 μ m.
Fig. 3 a is that to utilize diameter be the laser confocal microscope photo of the prepared PLLA porous support of the paraffin microballoon of 150～280 μ m.PLLA concentration 0.05g/mL, wax ball consumption 1.6g/mL polymers soln.
Fig. 3 b is that to utilize diameter be the laser confocal microscope photo of the prepared PLLA porous support of the paraffin microballoon of 280～450 μ m.PLLA concentration 0.05g/mL, wax ball consumption 1.6g/mL polymers soln.
Fig. 3 c is that to utilize diameter be the laser confocal microscope photo of the prepared PLLA porous support of the paraffin microballoon of 450～700 μ m.PLLA concentration 0.05g/mL, wax ball consumption 1.6g/mL polymers soln.
Fig. 4 a is PLLA concentration 0.04g/mL, the stereoscan photograph of prepared PLLA porous support under the condition of wax ball consumption 1.6g/mL polymers soln.Wax spherolite footpath: 150～450m.
Fig. 4 b is PLLA concentration 0.05g/mL, the stereoscan photograph of prepared PLLA porous support under the condition of wax ball consumption 1.6g/mL polymers soln.Wax spherolite footpath: 150～450 μ m.
Fig. 4 c is PLLA concentration 0.06g/mL, the stereoscan photograph of prepared PLLA porous support under the condition of wax ball consumption 1.3g/mL polymers soln.Microspherulite diameter: 150～450 μ m.
Embodiment
Can understand the present invention better by following embodiment, but these examples are not used for limiting the present invention.
Embodiment 1: the preparation of poly--L-lactic acid (PLLA) porous support
Adopt suspension method to prepare wax ball particulate.In an Erlenmeyer flask, adding 40g solid paraffin and 200mL concentration is the aqueous gelatin solution of 20mg/mL, is heated to 80 ℃, and paraffin becomes liquid state.Utilize magnetic stirrer to stir, stirring velocity is 200rpm, and the paraffin of liquid state is separated into small droplets.After treating paraffin suspension in stable condition, pour into rapidly in 0 ℃ the frozen water and carry out quenching, obtain solid paraffin microballoon.Adopt the domestic standard sieve that the paraffin microballoon is carried out classification, obtaining particle diameter is the paraffin microballoon of 150～450 μ m.
In a stainless steel mould, be the PLLA/ dioxane solution uniform mixing of 0.06g/mL with above-mentioned paraffin microballoon and concentration, the consumption of paraffin microballoon is every milliliter of PLLA solution of 1.3g.Said mixture is freezing under-25 ℃, PLLA solution is separated.Because the fusing point of dioxane is 10 ℃, mixture solidifies, and becomes solid simultaneously.Solid mixture is taken out from mould, and lyophilize was removed dioxane in 6 hours, obtained the mixture of paraffin microballoon and PLLA.Utilize the ebullient normal hexane to clean then, the extracting of paraffin microballoon is removed.The multiple said process of bob-weight is thoroughly to remove deparaffnize.Vacuum-drying finally obtains the PLLA porous support, as shown in Figure 1 to remove normal hexane.As can be seen from Figure, both having contained because of the paraffin microballoon is removed the diameter that produces in the porous support is macropore more than the 150 μ m, and also containing the diameter that produces because of the thermic phase separation in the inside of hole wall is the micropore of 20～30 μ m.Resulting porous support has good hole connectedness.
Embodiment 2: the control in poly--L-lactic acid (PLLA) porous support aperture
Prepare the paraffin microballoon according to the method in the application example 1, and utilize domestic standard sieve with the paraffin microballoon by size be divided into below the 150 μ m, not at the same level part of 150～280 μ m and 280～450 μ m, shown in Fig. 2 a, Fig. 2 b and Fig. 2 c.
Utilize the paraffin microballoon of above-mentioned different-grain diameter respectively, prepare the PLLA porous support according to the method among the embodiment 1, wherein the PLLA strength of solution is 0.05g/mL, and the consumption of paraffin microballoon is every milliliter of PLLA solution of 1.6g.The laser confocal microscope photo of resulting PLLA porous support is respectively shown in Fig. 3 a, Fig. 3 b and Fig. 3 c.The aperture of porous support can be controlled easily by the particle diameter of control paraffin microballoon as seen from Figure 3.
Embodiment 3: the poly--porosity of L-lactic acid (PLLA) porous support and the control of apparent density
Adopt the paraffin microballoon of different-grain diameter, different PLLA strength of solution and different paraffin microballoon consumptions respectively, and prepare the PLLA porous support according to the method for example 1.Table 1 has been summed up the porosity and the apparent density of PLLA porous support prepared under the various different conditions.As seen from Table 1, the particle diameter of the porosity of porous support and apparent density and paraffin microballoon is irrelevant, and is subjected to the influence of PLLA strength of solution and wax ball consumption bigger.Under the higher PLLA strength of solution, the apparent density of gained porous support is bigger, and porosity is lower; Paraffin microballoon consumption is big more, and the apparent density of gained support is more little, and porosity is high more.Fig. 4 a, Fig. 4 b and Fig. 4 c are the stereoscan photograph of the prepared PLLA porous support of PLLA solution and the different wax ball consumptions of different concns.Resulting PLLA porous support is fine and close when adopting higher PLLA concentration and lower wax ball consumption as seen from Figure 4, and resulting PLLA porous support is more loose when lower PLLA concentration of employing and higher wax ball consumption.
Table 1 is the apparent density and the hole counting rate meter 1 of PLLA porous support prepared under different wax spherolites footpath, PLLA concentration and the wax ball consumption
| Wax spherolite footpath, μ m | PLLA concentration, g/ml | Wax ball consumption, g/ml | Apparent density, g/cm 3 | Porosity, % |
| ??150～450 | ????0.05 | ????1.6 | ????0.023 | ????98.2 |
| ??150～450 | ????0.06 | ????1.6 | ????0.031 | ????97.6 |
| ??150～450 | ????0.07 | ????1.6 | ????0.038 | ????97.0 |
| ??150～280 | ????0.05 | ????1.3 | ????0.035 | ????97.2 |
| ??280～450 | ????0.05 | ????1.3 | ????0.032 | ????97.5 |
| ??450～710 | ????0.05 | ????1.3 | ????0.033 | ????97.4 |
| ??150～280 | ????0.05 | ????1.6 | ????0.021 | ????98.3 |
| ??280～450 | ????0.05 | ????1.6 | ????0.022 | ????98.3 |
| ??450～710 | ????0.05 | ????1.6 | ????0.022 | ????98.3 |
| ??150～280 | ????0.05 | ????2.0 | ????0.017 | ????98.7 |
| ??280～450 | ????0.05 | ????2.0 | ????0.019 | ????98.5 |
| ??450～710 | ????0.05 | ????2.0 | ????0.017 | ????98.7 |
Claims (4)
1. the paraffin microballons porous making agent method combines with thermally induced phase separation and prepares the method for used in tissue engineering porous polymer scaffold, it is characterized in that may further comprise the steps:
(1) adopt the suspended dispersed legal system to be equipped with the paraffin microballoon that particle diameter is 50 μ m～700 μ m;
(2) polymers soln that the paraffin microballoon that obtains and concentration are 0.01-0.5g/mL is under 15～55 ℃ of temperature, mixed by 0.5～3g paraffin microballoon/mL polymers soln is even, reduce temperature then to-100～10 ℃, polymers soln is separated;
(3) solvent in the polymers soln is removed in lyophilize, obtains paraffin microballoon and mixture of polymers;
(4) above-mentioned paraffin microballoon and mixture of polymers are placed the washing of ebullient normal hexane repeatedly, perhaps utilize cable-styled extractor that the extracting of paraffin microballoon is removed;
(5) normal hexane is removed in vacuum-drying, obtains porous polymer scaffold.
2. be separated with thermic by claims 1 described wax ball method pore-creating agent and combine the method for preparing the used in tissue engineering porous polymer scaffold, it is characterized in that said suspension method prepares the paraffin microballoon and is: with solid paraffin and concentration is the aqueous dispersant mixing of 1～100mg/mL, solid paraffin/aqueous dispersant ratio is 1～100g/100mL, be heated to 60 ℃～100 ℃, make paraffin become liquid state, stir, the paraffin of liquid state is separated into small droplets, after treating paraffin suspension in stable condition, pour quenching in-5 ℃～10 ℃ the cold water fast into, obtain solid paraffin microballoon.
3. be separated with thermic by claims 2 described wax ball method pore-creating agents and combine the method for preparing the used in tissue engineering porous polymer scaffold, it is characterized in that the dispersion agent that suspension method prepares the paraffin microballoon comprises gelatin, polyvinyl alcohol, polyoxyethylene glycol, sodium laurylsulfonate.
4. be separated with thermic by claims 1 described wax ball method pore-creating agent and combine the method for preparing the used in tissue engineering porous polymer scaffold, it is characterized in that said polymkeric substance comprises polyglycolic acid (PGA), lactic acid-ethanol copolymer (PLGA), polycaprolactone (PCL), urethane (PU), polycarbonate, poly-acid anhydrides, poly phosphazene, poly butyric ester and multipolymer thereof, collagen, chitosan etc. and above-mentioned mixture of polymers and the multipolymer between them, the product that also comprises the different spaces structure of above-mentioned polymkeric substance, as Poly-L-lactic acid (PLLA), dextrorotation poly(lactic acid) (PDLA) and meso poly(lactic acid) (PD, LLA).
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| CN102971018B (en) * | 2010-05-11 | 2016-02-03 | 阿勒根公司 | Porogen composition, Preparation method and use |
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| CN103269850B (en) * | 2010-09-09 | 2016-12-07 | 细胞吸附剂公司 | size selective polymer system |
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| CN101979102B (en) * | 2010-09-30 | 2013-03-13 | 中山大学 | Method and equipment for preparing tissue engineering scaffold with anisotropic pore structure |
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| CN102839433B (en) * | 2012-09-08 | 2015-04-29 | 福建师范大学 | Preparation method of polymer nanometer fiber bunch under independent state |
| CN104826163A (en) * | 2015-04-15 | 2015-08-12 | 赵红斌 | Composite scaffold material with effect of effective bone injury repair increase |
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| CN108553685A (en) * | 2018-04-09 | 2018-09-21 | 徐州医科大学 | The preparation method of artificial alveolar |
| CN111990477A (en) * | 2020-08-12 | 2020-11-27 | 华南理工大学 | Starch-based steady-state vegetable oil compound and preparation method thereof |
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