KR20190034123A - Non-solvent induced phase separation (NIPS)-based 3D plotting for porous scaffolds with core-shell structure - Google Patents

Abstract

The present invention relates to a method for preparing a porous support using a non-solvent phase separation method, which provides excellent stability and mechanical strength in vivo. The method of the present invention comprises a step of extruding a biodegradable polymer solution into a precipitation solution by using a 3D printer, wherein the precipitation solution is a mixed solution of water and alcohol.

Description

[0002] Non-solvent induced phase separation (NIPS) -based 3D plotting for porous scaffolds with core-shell structure,

TECHNICAL FIELD The present invention relates to a porous core-dense shell structured scaffold manufacturing technique using a non-solvent separation method.

Scaffolding made with 3D printers has the advantage of being able to provide that shape to the individual for the individual. In addition, the three dimensional macropore structure of the scaffold imparted through the 3D printer has the advantage of improving the regeneration speed of the tissue. On the other hand, the three-dimensionally connected micro-pore structure formed inside the scaffold is advantageous not only in the durability of tissue regeneration, which is similar to the pore structure of the living body, but also in induction / stimulation of cell adhesion, proliferation and differentiation.

1. Korean Patent Publication No. 10-2018-0001191

The present invention aims at producing a porous support (scaffold) having three-dimensional macroscopic pores using a biodegradable polymer having excellent biocompatibility and mechanical strength.

The present invention comprises a step of extruding a biodegradable polymer solution into a precipitating solution using a 3D printer,

Wherein the precipitating solution is a mixed solution of water and alcohol.

The present invention also relates to a porous support prepared by the above-

The filament constituting the porous support provides a porous support having a core of a porous structure and a structure of a shell of a dense structure.

The porous support according to the present invention can provide stability in vivo by using a biodegradable polymer having excellent biocompatibility. Further, since the biodegradable polymer has excellent mechanical strength, it can provide excellent mechanical strength in vivo have.

The porous support according to the present invention has a structure in which the filaments constituting the porous support have a porous core structure and a shell having a dense structure. Such a structure has an internal structure similar to that of a living bone, Can induce / stimulate not only excellent mechanical strength but also cell attachment, proliferation and differentiation.

The porous support according to the present invention has a dense structure of the shell (outer portion) of the filament, unlike other conventional techniques, so that the outer portion of the support may have a dense structure without pores.

Further, in the present invention, the thickness and thickness of the shell portion of the filament can be controlled by adjusting the composition and the ratio of the precipitating solution, thereby controlling the strength of the support artificially.

FIG. 1 is a schematic view showing a method for producing a porous support according to the present invention. After a polycaprolactone solution is transferred to a piston, a scaffold is prepared in a precipitating solution according to a predetermined design using a 3D printer.
Fig. 2 is a schematic diagram showing a state in which a solution of polycaprolactone extruded from a piston enters a precipitating solution. Fig. When the solution of the polycaprolactone is exposed to the precipitating solution, the filament surface is laminated according to a predetermined design, and the surface of the filament becomes a solidified and pore structure through phase separation.
3 is a schematic view showing a cross section of a polycaprolactone filament according to the constitution of a precipitating solution.
4 is a schematic view showing a structure of a polycaprolactone filament, that is, a polycaprolactone support (scaffold) according to the constitution of a precipitating solution.
5 is an image showing a structure of a polycaprolactone filament, that is, a polycaprolactone support (scaffold) according to the constitution of a precipitating solution.
Figure 6 is an image of a polycaprolactone support (scaffold) prepared according to the proportions of water and ethanol in the precipitation solution.
7 is a photomicrograph showing the change in the pore structure of the produced polycaprolactone filament according to the ratio of water and ethanol in the precipitating solution.
8 is an enlarged photomicrograph showing the change in the pore structure of the produced polycaprolactone filament according to the ratio of water and ethanol in the precipitating solution.
9 is a photomicrograph showing the core pore structure of the produced polycaprolactone filament according to the ratio of water and ethanol in the precipitating solution.
10 is a graph showing the tensile strength of the prepared polycaprolactone scaffold (support) according to the ratio of water and ethanol in the precipitating solution.

The present invention comprises a step of extruding a biodegradable polymer solution into a precipitating solution using a 3D printer,

Wherein the precipitating solution is a mixed solution of water and alcohol.

Hereinafter, the porous support according to the present invention will be described in more detail.

In the present invention, a biodegradable polymer solution extruded into a precipitating solution by using a 3D printer can be expressed as a filament, and the laminate (structure, laminate) of the filaments can be expressed as a support or a scaffold.

The method for preparing a porous support according to the present invention includes a step of extruding a biodegradable polymer solution into a precipitating solution using a 3D printer.

In one embodiment, the method of the present invention comprises the steps of: preparing a biodegradable polymer solution;

Extruding a biodegradable polymer solution in a 3D printer capable of a three-dimensional lamination system and laminating the biodegradable polymer solution in a precipitation solution;

Placing the laminated filaments, that is, the porous support, in the precipitating solution; And

And removing the residual solvent and the precipitating solution from the porous support.

First, a biodegradable polymer solution is prepared by dissolving a biodegradable polymer in a solvent.

The type of the biodegradable polymer is not particularly limited and includes polycaprolactone (PCL), polylactide (PLA), polylactide glycolide (PLGA), polyglycolide , PGA), or a mixture thereof.

The solvent of the biodegradable polymer solution is not particularly limited as long as the biodegradable polymer can be dissolved. For example, dichloroethane (DCE), tetrahydrofuran (THF), chloroform, acetone acetone, methylethylketone (MEK), and hexafluoro-2-propanol (HFP) may be used.

The concentration of the biodegradable polymer in the biodegradable polymer solution may be 12 to 25 wt%. And may have a suitable viscosity to laminate the filaments using the 3D printer in the above range. If the content of the biodegradable polymer is less than 12 wt%, the extruded filament is difficult to have a uniform shape and may collapse in the vertical direction. When the amount exceeds 25 wt%, the biodegradable polymer is hardly dissolved in the solvent and the viscosity is rapidly increased, There is a possibility that it is difficult to extrude from the nozzle tip of the syringe.

In the present invention, in order to increase the solubility of the biodegradable polymer, the biodegradable polymer and the solvent may be further stirred at 40 to 70 ° C or 50 to 60 ° C. The stirring may be carried out by a stirring method such as a magnetic stirring method, a mechanical stirring method or a vortex method commonly used in the art. After the stirring is completed, the mixture may be cooled to room temperature and used in the next step.

In the next step, a biodegradable polymer solution is extruded from a 3D printer capable of a three-dimensional lamination system and laminated in a precipitation solution.

In this step, the porous support having controlled pore and pore structures of various sizes in the polymer skeleton forming the porous structure can be prepared by laminating the biodegradable polymer solution three-dimensionally in the precipitating solution.

At this time, the precipitating solution is a non-solvent, specifically, a mixed solution of water and alcohol. As the alcohol, ethanol, methanol, butanol, propanol or a mixture thereof can be used. The ratio of water and alcohol in the precipitation solution may be 100: 10 to 500, 100: 10 to 300, or 100: 10 to 100. The shell portion of the filament produced in the above range may have a dense structure.

In the present invention, the biodegradable polymer solution is phase-separated by phase exchange between the solvent of the biodegradable polymer solution and the non-solvent of the precipitation solution, and the biodegradable polymer solution is coagulated and laminated, and a pore structure is formed in the filament. The pores may have a micro-size.

In one embodiment, the extrusion and lamination of the biodegradable polymer solution is performed using a 3D printer, which allows the user to program or laminate the desired structure by using the 3D printer. The biodegradable polymer solution can be extruded into the precipitation solution along with the coordinates of the pre-designed 3D printer, and a support having a complicated structure can be easily manufactured. The speed of the printer during lamination can be controlled according to the extrusion speed. The 3D printer is connected to an extrusion system (extruder) for extruding filaments, and the extruded filaments can be laminated according to a programmed design. Specifically, the extruded filaments can be laminated along a desired direction and laminated a plurality of times.

Such a laminate can be laminated in three dimensions, and in one embodiment, the filaments can be laminated to have unidirectional properties. The unidirectional means that the filaments are oriented in one direction with respect to one layer of filaments stacked at regular intervals in a filament support having a plurality of filaments. That is, when the filaments are formed in multiple layers, one layer is formed such that the filaments are spaced apart in one direction, and the other layer in contact with the one layer is formed such that the filaments are spaced apart in a direction perpendicular to one layer . At this time, the spacing between the filaments may be 10 to 1000 mu m.

In order to induce proper solidification of the biodegradable polymer solution in the three-dimensional lamination, the extrusion rate is preferably 1 to 5 mm / s. The extrusion rate means the rate of lamination of the filaments and the rate of phase exchange between solvent and non-solvent. For example, when the extrusion speed is less than 1 mm / s, there is a possibility that the upper filament collapses in the direction of gravity between the already laminated filaments and the filaments during the three-dimensional lamination, There is a possibility that the piling structure collapses in the vertical direction because the speed of the lamination is higher than the speed at which the polymer solution is solidified.

The compressed air during the lamination may be treated at 10 to 35 kPa. As the concentration of the biodegradable polymer solution increases, the viscosity also increases, so it is preferable to adjust the compressed air to the range as shown for accurate lamination.

The diameter of the nozzle during extrusion may be 1,000 占 퐉, 800 占 퐉 or 600 占 퐉 or less.

The next step is to place the stacked filaments, that is, the porous support, in the precipitation solution.

Through this step, complete phase separation between the solvent of the biodegradable polymer solution and the non-solvent of the precipitation solution is possible.

In this step, the standing time may be 1 to 30 minutes.

In the present invention, a step of removing solvent, precipitating solution, impurities and the like from the porous support is further performed.

Examples of the method for removing the solvent and the like include freeze drying, vacuum drying, natural drying, and the like, and may include an additional washing step.

In the case of lyophilization, it can be carried out at a temperature of -80 to -10 ° C, specifically -50 to -20 ° C, and the freezing medium can be easily removed without damaging the laminate of the molded body in this temperature range.

By the manufacturing method according to the present invention, a porous support having filaments aligned in one direction is formed. The unidirectional means that the filament is oriented in one direction with respect to one layer in which the filaments are stacked at a predetermined interval in the porous support in which filaments are formed in multiple layers.

The present invention also relates to a porous support produced by the above-mentioned production method.

In one embodiment, the porous support has a structure in which filaments having an average diameter (diameter) of 300 to 1000 mu m, 400 to 900 mu m, or 500 to 800 mu m are formed in one direction for each layer.

Macropores having an average diameter (diameter) of 10 to 1000 mu m or 100 to 900 mu m or 200 to 800 mu m are formed between the filaments of the porous support.

Particularly, in the porous support, the filament constituting the support has a structure of a porous structure core and a dense structure shell. Here, the core and the shell are represented by a shell, which surrounds the inside where the pores are formed and the inside where the pores are formed, with reference to a section perpendicular to the direction in which the parentheses are extruded. The ratio of the shell to the filament may be from 1 to 50%.

This is due to the constitution of the precipitate solution in the production process. When alcohol is used as the precipitating solution, micro- or nanoporous filaments are formed. However, when a mixed solution of water and alcohol is used as in the present invention, filaments having pores formed in the core and a dense structure in the shell portion, that is, filaments having two structures, can be produced.

In the present invention, the core has pores of 10 to 500 nm, 20 to 100 nm or 30 to 50 nm.

That is, the porous support according to the present invention has a micro pore structure and a macro pore structure.

The present invention also relates to a product comprising the porous support.

Since the porous support according to the present invention has a microporous / macropore double pore structure and excellent physical properties, it can be used not only for regenerating hard tissues such as bones and teeth of a human body but also for regenerating various soft tissues such as cartilage, skin, ligament and muscle .

In the present invention, a porous support having a high porosity and a three-dimensionally connected pore structure is prepared through the exchange of a solvent and a non-solvent by using a non-solvent separation method for producing a porous support. The porous support prepared by the non-solvent separation method may have a pore structure having a structure similar to a living bone.

The present invention is different from the conventional art in that a shell portion densified without pore structure can be artificially provided by controlling the type and content ratio of the precipitating solution. The densified shell can easily adjust the thickness of the shell by changing the composition of the precipitating solution, and the mechanical strength of the support can be controlled by adjusting the thickness of the shell.

Advantages and features of the present invention and methods of achieving them will become apparent with reference to the embodiments described in detail below. The present invention may, however, be embodied in many different forms and should not be construed as being limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art. Is provided to fully convey the scope of the invention to those skilled in the art, and the invention is only defined by the scope of the claims.

Example

Example  1. Core-Shell Scaffold  Produce

(One) Scaffold  Manufacture of solutions for production

10 g of polycaprolactone (Sigma-Aldrich, St. Louis, Mo., USA) and 57.4 ml of acetone were placed in a 100 ml sealable glass bottle, and the mixture was stirred at 50 ° C for 2 hours in an agitator to dissolve the polycaprolactone in acetone A fully dissolved polycaprolactone solution was prepared. The polycaprolactone solution was cooled at room temperature for 2 hours.

(2) Through the sale of revenue Scaffold  making

In order to prepare a predetermined pore shape, a set amount of ethanol and water was mixed to prepare a precipitate solution.

The polycaprolactone solution prepared in (1) above was charged into a 15 ml extruder piston having a nozzle diameter of 0.5 mm.

The polycaprolactone solution contained in the extrusion piston was connected to a 3D printer (Ez-Robo 5), and a pressure of 25 kgPa was applied at a rate of 3 mm per second at a lamination speed of 11.6 mm x 11.6 mm x 1.3 mm) was prepared in the precipitation solution (see Fig. 1).

After the polycaprolactone scaffold was completed in the precipitation solution, the scaffold was exposed for 5 minutes in the precipitation solution for complete phase separation.

(3) Scaffold  Solvent and Expense  remove

The scaffold having been completely phase-separated by the above step (2) was washed with 200 ml of distilled water for 5 minutes and lyophilized for 24 hours to completely remove ethanol and distilled water remaining on the spherical supporter.

Experimental Example  1. Configuration change of filament according to the composition of precipitate solution

2 is a schematic diagram showing a state in which a solution of a polycaprolactone extruded from a piston enters a precipitating solution. When the solution of the polycaprolactone is exposed to the precipitating solution, the filaments are laminated according to a preset design, and the surface of the filament becomes a solidified and pore structure through phase separation.

In the present invention, in order to confirm the compositional change of the filament according to the constitution of the precipitating solution, the filament was extruded into the precipitating solution while changing the ratio of water and ethanol constituting the precipitating solution.

3 is a schematic view showing a cross section of a polycaprolactone filament according to the constitution of a precipitating solution.

As shown in FIG. 3, in the precipitate solution of 100% ethanol, the filament shows a structure without a dense shell. As the content of water increased, a filament having a core having a porous structure and a shell having a dense structure was produced.

That is, as the content of water in the precipitate increases, the shell part becomes thicker.

FIG. 4 is a schematic view showing a structure of a polycaprolactone filament according to the constitution of a precipitating solution, that is, a scaffold, and FIG. 5 is an image of the scaffold.

As shown in FIG. 4, in the precipitate solution of 100% ethanol, only a pore structure having no shell is formed. In the precipitate solution of 100% of water, it can be seen that the porous structure core and the shell structure of a dense structure are formed.

Further, as shown in the image of FIG. 5, it can be confirmed that the polycaprolactone filaments are laminated regardless of the precipitating solution to produce a scaffold. As in the schematic diagram of FIG. 4, the scaffold produced by the present invention has a shell-free pore structure in the precipitate solution of 100% ethanol. In the precipitate solution of 100% of water, the core of the porous structure and the shell of the dense structure Structure.

Experimental Example  2. Constitutional change of filament according to the ratio of water and ethanol in the precipitate solution

The scaffold was prepared by changing the ratio of water and ethanol in the precipitation solution as shown in Table 1 below. The preparation of the scaffold was made by the method of Example.

The ratio of water to ethanol in the precipitation solution A 0 (water): 100 (ethanol) B 25: 75 C 50: 50 D 75: 25 E 100: 0

Figure 6 is an image of a polycaprolactone scaffold prepared according to the proportions of water and ethanol in the precipitation solution.

The size of the scaffold is 11.6 mm x 11.6 mm x 1.3 mm, and it can be confirmed that the scaffold can be manufactured regardless of the composition and the ratio of the precipitating solution.

FIG. 7 is a micrograph showing a change in the pore structure of the produced polycaprolactone according to the ratio of water and ethanol in the precipitating solution, and FIG. 8 is a micrograph further enlarged. The thickness of the shell having a dense structure is shown in Table 2 below.

The ratio of water to ethanol in the precipitation solution Thickness of shell part (μm) A 0 (water): 100 (ethanol) 0 B 25: 75 9.7 C 50: 50 15.3 D 75: 25 24.9 E 100: 0 34.0

As shown in FIGS. 7 and 8, as the ratio of water in the precipitate increases, the shell portion of the dense structure increases, which can be confirmed from the increase in thickness of the shell portion as shown in Table 2.

9 is a photomicrograph showing the core pore structure of the produced polycaprolactone according to the ratio of water and ethanol in the precipitation solution.

As shown in FIG. 9, it can be seen that the pore structure of the core portion has a constant structure irrespective of the constitution of the precipitating solution.

Experimental Example  3. The ratio of water and ethanol in the precipitation solution Scaffold  Tensile strength measurement

The tensile strength of the scaffold used in Experimental Example 2 was measured and shown in Fig. 10 and Table 3 below.

The ratio of water to ethanol in the precipitation solution Tensile Strength (MPa) A 0 (water): 100 (ethanol) 0.83 B 25: 75 1.04 C 50: 50 1.17 D 75: 25 1.25 E 100: 0 1.39

As shown in FIG. 10 and Table 3, it can be seen that as the content of water in the precipitate increases, that is, as the thickness of the shell portion of the dense structure increases, the tensile strength increases proportionally.

Claims (8)

And extruding the biodegradable polymer solution into the precipitating solution using a 3D printer,
Wherein the precipitating solution is a mixed solution of water and alcohol.
The method according to claim 1,
The biodegradable polymer may be selected from the group consisting of polycaprolactone (PCL), polylactide (PLA), polylactide glycolide (PLGA), polyglycolide (PGA) A method for producing a porous support.
The method according to claim 1,
The solvent of the biodegradable polymer solution is dichloroethane (DCE), tetrahydrofuran (THF), choloroform, acetone, methylethylketone (MEK) and hexafluoro-2-propanol (hexafluoro-2-propano, HFP).
The method according to claim 1,
Wherein the alcohol in the precipitation solution is ethanol, methanol, butanol, propane or a mixture thereof.
The method according to claim 1,
Wherein the ratio of water to alcohol in the precipitating solution is 100: 10 to 500. < Desc / Clms Page number 24 >
The method according to claim 1,
And allowing the support extruded in the precipitation solution to stand in the precipitation solution for 1 to 30 minutes.
A process for the production of a compound according to claim 1,
The filament constituting the porous support has a core of a porous structure and a structure of a shell of a dense structure.
8. The method of claim 7,
The percentage of shell in the filaments is 1 to 50%.

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