Title:  Preparing porous biodegradable polymeric scaffolds for tissue engineering using effervescent salts

United States Patent:  6,586,246

Issued:  July 1, 2003

Inventors:  Yoon; Jun-Jin (Seoul, KR); Park; Tae-Gwan (Daejeon, KR); Nam; Yoon-Sung (Seoul, KR)

Assignee:  Innotech Medical, Inc. (Daejon, KR)

Appl. No.:  700458

Filed:  February 6, 2001

PCT Filed:  February 12, 2000

PCT NO:  PCT/KR00/00106

PCT PUB.NO.:  WO00/55300

PCT PUB. Date:  September 21, 2000

Abstract

Three-dimensional porous biodegradable and biocompatible, polymeric scaffolds for tissue engineering are prepared by a method involving effervescing of an effervescent salt in a gel to result in a porous structure. A polymer is dissolved in an organic solvent to prepare a polymer solution of high viscosity. Optionally, the polymer solution is mixed with an organic solvent that does not dissolve the polymer to concentrate the solution. An effervescent salt is homogeneously mixed with the polymer solution to give a polymer/salt/organic solvent mixed gel. The organic solvent is removed from the mixed gel to produce an organic solvent-free polymer/salt gel slurry. The gel slurry is submerged in a hot aqueous solution or acidic solution to cause the salt to effervesce at room temperature to form a porous three-dimensional polymeric structure. The polymeric structure is washed with distilled water and freeze-dried to yield a scaffold that is suitable for cell or tissue culture. Pore size and porosity of the scaffold can be easily controlled by controlling the size and amount of the effervescent salt and concentration of the acidic solution.

DISCLOSURE OF THE INVENTION

It is an object of the present invention to overcome the above problems encountered in prior arts and to provide a method for preparing biodegradable, three-dimensional, porous scaffolds for tissue culture, whereby the scaffolds can be molded in desirable forms and have desirable pore sizes and porosities.

Based on the present invention, the above object could be accomplished by a provision of a method for preparing biodegradable, three-dimensional, porous scaffolds for tissue culture, comprising the steps of dissolving a biodegradable polymer in an organic solvent to prepare a polymeric solution of high viscosity, homogeneously mixing an effervescent salt in the polymeric solution to give a polymer/salt/organic solvent mixed gel, removing the organic solvent from the polymer/salt/organic solvent mixed gel, submerging the organic solvent-free polymer/salt gel slurry in a hot aqueous solution or acidic solution to render the salt to effervesce at room temperature to afford a three-dimensional polymeric structure, and washing with distilled water and freeze-drying the polymeric structure.

BEST MODES FOR CARRYING OUT THE INVENTION

Before the present method for preparing (title of the invention) are disclosed or described, it is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting. It must be noted that, as used in the specification and the appended claims, the singular forms "a", "an" and "the" include plural referents unless the context clearly dictates otherwise.

Throughout this application, where publications are referenced, the disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains.

In the present invention, the preparation of biodegradable and biocompatible, three-dimensional, porous scaffolds for tissue culture is based on phase separation and particulate leaching. First, a biodegradable and biocompatible polymer is dissolved in an inorganic solvent to give a highly concentrated solution with high viscosity. Optionally, the polymer solution is further mixed with an organic solvent which does not dissolve the polymer, so as to concentrate the solution into a gel phase of a concentrated solution. By doing this, it is possible to prepare porous polymeric scaffolds even with biodegradable, low-molecular weight polymers which cannot be conventionally used as materials on account that their solutions are of low viscosity even at high concentrations. Then, the polymeric solution is homogeneously mixed with an effervescent salt, followed by the removal of the organic solvent from the resulting polymer/salt/organic solvent mixed gel. Submergence of the organic solvent-free polymeric/salt gel slurry in a hot aqueous solution or acidic solution effervesces the salt, resulting in a porous structure. Subsequently, this structure is washed with distilled water and freeze-dried to yield a scaffold, which is suitable for cell or tissue culture.

Useful as the biodegradable and biocompatible polymer in the present invention is one selected from the group consisting of poly(L-lactic acid) (PLLA), amorphous poly(D,L-lactic acid) (PDLLA), poly(glycolic acid), poly(D,L-lactic-co-glycolic acid) (PLGA), poly(caprolactone), poly(hydroxy butyrate), and copolymers of these polymers. They may be used irrespective of molecular weight, but better results are obtained from those whose molecular weight is in the range of 5,000-500,000.

Examples of the organic solvent for use in dissolving the biodegradable polymers include methylene chloride, chloroform, acetone, dimethylsulfoxide, dimethylformamide, N-methylpyrrolidone, dioxane, tetrahydrofluran, ethylacetate, methylethylketone, and acetonitrile. Incapable of dissolving the biodegradable polymer, the organic solvent used for the concentration of the polymeric solution is exemplified by ethanol, methanol, aqueous ethanol, ethyl ether, diethyl ether, hexane, petroleum ether, and aqueous petroleum ether.

With a size of 100-500 .mu.m, the effervescent salt is selected from the group consisting of ammonium carbonate, ammonium bicarbonate, sodium carbonate, and sodium bicarbonate. It is preferred to use the salt at such an amount that the weight ratio of the polymer to the effervescent salt may be in the range from 1:1 to 1:100.

Depending on the organic solvent remaining in the polymer/salt/organic solvent mixed gel, various methods may be utilized to remove the organic solvent. Organic solvents with relatively low boiling points, such as methylene chloride, chloroform and dioxane, are removed through drying at ordinary pressure or under vacuum while high-boiling point solvents, such as dimethylsulfoxide (DMSO) and methylpyrrolidone, are replaced with low boiling point solvents, such as ethanol and methanol, before being dried at atmospheric pressure or under vacuum.

In order to effervesce the salt of the polymer/effervescent salt slurry, hot distilled water or acidic aqueous solutions are used. It takes a lengthy period of time to conduct the effervescence of the salt with tepid, distilled water. If the temperature of the distilled water is raised, the reaction time can be reduced. The temperature needs not to be particularly limited, but may be selected according to the judgement of those who are skilled in the art. On the other hand, acidic aqueous solutions enable the salt to effervesce at room temperature within a short period of time. In addition, their concentrations affect the size of the pores formed in the scaffolds, so that the pore size can be under the control of the concentration. Therefore, the effervescence of the salt with the acidic aqueous solutions makes it possible to prevent the thermal distortion of the polymer and to form pores at desirable sizes as well as to settle down inside the porous scaffold the drugs necessary for cell culture.

Available in the effervescence of the present invention is the acid selected from the group consisting of citric acid, hydrochloric acid, acetic acid, formic acid, tartaric acid, salicylic acid, benzoic acid, and glutamic acid. For use, these acids are dissolved to the concentration of 1% or supersaturation in water or in an aqueous solution saturated with an organic solvent, such as methylene chloride, chloroform, dioxane, dimethylsulfoxide (DMSO), and methyl pyrrolidone.

Claim 1 of 12 Claims

What is claimed is:

1. A method for preparing a biodegradable and biocompatible, porous, polymeric scaffold for tissue engineering, comprising the steps of:

dissolving a biodegradable polymer in an organic solvent to prepare a polymeric solution of high viscosity;

homogeneously mixing an effervescent salt in the polymeric solution to give a polymer/salt/organic solvent mixed gel;

removing the organic solvent from the polymer/salt/organic solvent mixed gel to produce an organic solvent-free polymer/salt gel slurry;

submerging the organic solvent-free polymer/salt gel slurry in a hot aqueous solution or acidic solution to cause the salt to effervesce at room temperature to form a three-dimensional polymeric structure; and

washing the three-dimensional polymeric structure with distilled water and freeze-drying the washed polymeric structure.

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