CA1170001A - Method for preserving porosity in porous materials - Google Patents
Method for preserving porosity in porous materialsInfo
- Publication number
- CA1170001A CA1170001A CA000381746A CA381746A CA1170001A CA 1170001 A CA1170001 A CA 1170001A CA 000381746 A CA000381746 A CA 000381746A CA 381746 A CA381746 A CA 381746A CA 1170001 A CA1170001 A CA 1170001A
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- Prior art keywords
- highly porous
- porous foam
- porosity
- collapsible
- dimensionally
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- Manufacture Of Porous Articles, And Recovery And Treatment Of Waste Products (AREA)
- Materials For Medical Uses (AREA)
Abstract
ABSTRACT
A dimensionally-stable, non-collapsible highly porous foam based upon an insoluble protein-based polymeric material, is produced by forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated tempera-tures, quickly freezing the dispersion to form frozen liquid particles, subliming the frozen liquid particles to produce a highly porous foam, and subjecting the highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize the highly porous foam so that its porosity is preserved when it is contacted with a liquid solution of a chemical cross-linking agent. The stabilized highly porous foam is then subjected to a liquid solution of a chemical crosslinking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
A dimensionally-stable, non-collapsible highly porous foam based upon an insoluble protein-based polymeric material, is produced by forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated tempera-tures, quickly freezing the dispersion to form frozen liquid particles, subliming the frozen liquid particles to produce a highly porous foam, and subjecting the highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize the highly porous foam so that its porosity is preserved when it is contacted with a liquid solution of a chemical cross-linking agent. The stabilized highly porous foam is then subjected to a liquid solution of a chemical crosslinking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
Description
~ 3~00n~
Description MEI'HOD FOR PRESERVING POROSITY IN POROUS MATERIALS
Government Support The inventiorl described herein was made in the course of or under grants from the National Institutes of Health.
Field of_the Invention This invention is in the field of materials and more particularly relates to the treatment of porous materials to preserve their porosity.
Backqround of the invention Many materials are formed to contain a large number of poresO For example, certain materials or non-woven materials are porous, as are a large number of foams based upon natural or synthetic polymers. In many applications for such porous materials, it is important for the porous material to retain its porosity during exposure of the material to liquids without collapse of the pores.
One example of materials where this is true is a new class of tissue-compatible materials which are also insoluble in the presence of body fluids and controllably degradable in the presence of body enzymes and has been disclosed in U.S. Patent No. 4,280,954.
These materials are known as crosslinked collagen-mucopolysaccharide composites. They are synthe-sized by intimately contacting collagen with a muco-polysaccharide and subsequently crosslinking the resulting productO Suitable collagens can be derived .
1 1 ~0 ~
from a number of animal sources, and suitable mucopoly-saccharides include, but are not limited to, chondroi-tin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate, keratan sulfate, heparan sulfate, heparin, and hyalu ronic acid. Insolublization can be achieved by chemi-cal, radiation, or other suitable crosslinking tech-niques, or dehydrothermal treatment. Dehydrothermal treatment is particularly preferred and is achieved by reducing the moisture level of the composites to a very low level, such as by subjecting the composite material to elevated temperatures and high vacuum.
~ hese crosslinked collagen-mucopolysaccharide com posites are believed to be comprised of collagen mole-cules or collagen fibrils with long mucopolysaccharide chains attached to them. Crosslinking appears to an-chor the mucopolysaccharide chains to the collagen so they will not elute or otherwise become disengaged.
These crosslinked collagen-mucopolysaccharide com-posites have been found to retain the advantageous properties of native collagen. Additionally, such ma-terials can be synthesized to be very weakly antigenic, degradable by collagenase or other enzymes at controll-able rates, and insoluble in the presence of body fluids. Additionally, such composites can be synthe-sized to have ultimate tensile strengths, elongationsat break, and other mechanical properties particularly desired for artificial skin grafts and wound dressings.
1 ~J~
These crosslinked collagen-mucopolysaccharide compos;tes have been combined with moisture trans-mission control layers adherently bonded thereto to form synthetic skin. The moisture transmission con-trol layers are formed from nontoxic materials whichcontrol moisture flux of the overall membrane and can be formed from synthetic polymers such as silicone resins, polyacrylate or polymethacrylate esters or their copolymers, and polyurethanes. Such synthetic skin is described in U.S. Patent No. 4,060,081, issued to Yannas et al. on November 29, 19770 In many applications for the crosslinked collagen-mucopolysaccharide composites, it has been found highly preferable to prepare these materials as highly porous foams. For example, in the use of these materials as medical prostheses, including their use as synthetic skin, there is frequently a need for migration of cells from adjacent host tissue into tha prosthesis.
Such migration provides firmer attachment of the prostheses to the tissue and is also indispensable whenever there is a requirement for the invading cells to synthesize new functional tissue inside the pores of the prosthesis, which, if biodegradable, eventually disappears from the original site leaving in its place the newly synthesized tissue. In short, it has been found that high porosity in these composite materials, very oten in excess of 90% pore volume, allows a significantly greater degree of cell infiltration, elicits a much reduced fibrous sac, and allows desired tissue synthesis to occur at a much faster rate than corresponding material produced as membranes without high porosity.
., ,."~
~ ~ ~looo ~
Despite the need for porous materials, most tradi~
tional technqiues for producing foams are not suitable to produce materials which are biocompatible. For ex-ample, it is well known to produce foamed solids by em-ploying blowing agents to produce synthetic polymericfoams. An example of the use of blowing agents is the formation of polyurethane foams. However, the prepara-tion of such foamed polyurethanes generally involves the use of toxic chemicals, such as diisocyanates, which may often remain in an incompletely reacted form. This would be particularly objectionable in the case of a collagen or collagen-mucopolysaccharide based material, which would evoke little or no inflammatory response itself, but would generate a toxic inflamma-tory response if unreacted diisocyante were present.
It is also difficult to control foam density or poros-ity using blowing agents, and even when possible it re-quires elaborate processing steps including addition of catalytic systems which also may be toxic.
Because of the problems with conventional foam generation techniques, it has been found preferable to produce porous materials based upon crosslinked colla-gen mucopolysaccharide composites by a technique known as freeze drying. In this technique, an aqueous dis-persion of the composite is quickly frozen and the re-sulting ice particles are subsequently caused to sub-lime in the presence of vacuum. A solid, highly porous material results, and the degree of porosity can be controlled by adjusting the concentration of solids in the dispersion prior to the rapid freezing, as well as by adjusting the temperature and vacuum to which the drying membrane is exposed during the process.
Although the freeze drying technique has been found to be generally suitable, the materials produced do suffer one disadvantage. This disadvantage is that the dry porous solids obtained by sublimation of ice under vacuum often shrink considerably and irreversibly when brought into contact with liquids, such as an aqueous solution. Such shrinkage causes closure of the pores and makes the material less useful in the appli-cations where the high level of porosity is required or preferable. Unfortunately, most applications for these porous biocompatible materials require that the mate-rial be further processed or stored in aqueous solu-tions, or require that the materials be placed in con-tact with aqueous body fluids during use which would also cause undesirable shrinkage of a prosthesis formed from such materials.
The problems of shrinkage and pore collapse suf-fered by porous crosslinked collagen-mucopolysaccharide materials are illustrative of problems encountered with porous materials based upon a wide variety of natural and synthetic polymers when such porous materials are brought into contact with liquids.
Disclosure of the Invention This invention relates to the treatment of porous materials, particularly highly porous materials (e.g., above 90% pore volume) with a combirlation of elevated temperature and vacuum to modify such materials so that their porosity ls preserved when they are subsequently contacted with fluids.
, . . .
0 1~ ~
More particularly, the invention provides a method of producing a dimensionally-stable, non-collapsible highly porous foam based upon insoluble protein-based polymeric material, which comprises the steps of:
a~ forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated temperatures;
b) quickly freezing the dispersion to form frozen liquid particles;
c) subliming the frozen liquid particles to produce a highly porous foam;
d) subjecting the highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize the highly porous foam so th~t its porosity is pre-served when it is contacted with a liquid solution of a chemical cross-linking agent, and, e) subjecting the stabilized highly porous foam to a liquid solution of a chemical cross-linking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
An irnportant advantage is the degree of control over the porosity of the resulting protein foams which can be gained. Porosity, for example, can be con-trolled by adjustment of the solids conten~ of the dis~
persion prior to the quick freezing step as well as by adjustment of the temperature and pressure employed during the freeze-drying process.
Porosity is preserved, even upon contact with liquids, by the treatment under elevated temperature and vacuurn. This prevents shrinkage of the foam materials when they are subsequently conkacted with liquids during further processing, storage or use.
This is especially advantageous since the most cornmon methods of cross]inking collagen involve contacting it with an aqueous solution of a crosslinking agent, such as glutaraldehyde. Without prior dehydrothermal treatment, it would be very difficult to crosslink , the collagen without damaging its porosity. In addition, one preferred method of storing the collagen membrane described in U.S. Patent 4,060,081 (Yannas et al, 1977) is in the hydrated form, in a sterile watertight pouch that contains a piece of membrane immersed in a saline solution.
One class of materials suitable for production of porous protein foams is the class of crosslin~ed colla-gen-mucopolysaccharide composite materials described above. However, protein foams can be formed from pure collagen or other proteins in their pure form, such as gelatin, albumin, fibrinogen, and soybean protein~
Additionally, collagen or these other protein molecules can be grafted with various comonomers to form composite or grafted protein foams.
The initial step in preparation of these protein foams is the formation of an aqueous dispersion of the protein. Collagen based material can be employed by first swelling the collagen in aqueous acidic medium.
In this regard, it is particularly preferred to swell collagen at a low p~, such as about 3.5 or lower. See, examples 16 and 17 of U.S~ Patent No. 4,280,954.
Control over porosity of the foams can be achieved at the dispersion ~tage by adjusting the solid content of the dispersion~ ~n general, higher porosity will be attained as the solids content is lowered. Addition-ally, control of porosity can be obtained by adjusting the temperature and vacuum during freeze-drying.
In regard to producing shaped articles from the dispersions of protein, there are several alternative methods which mày be employed. In one method, the aqueous dispersion is simply filtered to produce porous sheets which can subse~uently be shaped. On the other hand, more intricate shapes, such as are often required to produce arterial or benous tubing, can be produced by a crossflow filtration molding method described in U.S. Patent No. 4,252,759.
. ' .
1 ~ ~00~ ~, Porosity is achieved by freeze drying techniques which generally involve subjecting the shaped article to low temperatures, so that ice crystals are formed, followed by sublimation of the ice under vacuum. Such techniques are known to those skilled in the art and many of these have been described in the patent litera-ture. See, for example, U. S. Patents 3,632,371;
3,471,598; 3r368,911; and 2,610,625.
The shaped protein foams are treated under high temperature and vacuum. In general, temperatures of 80C to 180C have been found suitable for collagen based foams. The vacuum may vary from about 1 mtorr up to slight vacuum just below atmospheric pressure.
Practical combinations of temperatures and pressures for collagen based materials have been found to be 80C
at 50 mtorr, 105C at 1 torr and 150C at 600 torr~ An increase in temperature or vacuum can be used singly or in combination to accelerate the process of pore pre-servationO
As mentioned above, a particularly important use for the highly porous protein foams produced by the method described herein i5 in multilayer membranes use-ful to cover wound dressing. Thus, wo~nd dressings or synthetic skin can be produced by multilayer membrane formed from either collagen or crosslinked collagen-mucopolysaccharide composite used in conjunction with moisture transmisslon control layers as described in U. S. 4,060,081.
When used as synthetic skin, the multilayer 3a membrane described in U. S. 4,060,081 exhibits two very beneficial characteristics that result from very high porosity. First, if the porosity of the collagen material exceeds about 90%, and is preferably about 95% t~ 98%, then epithelial cells tend to grow between the collagen layer and the silicone layer.
~l~oonl .
Since the collagen layex must be covered ~y epithelial cells before it can be biodegraded and the wound fully closed, this is very desirable. Second, if the porosity is sufficiently high to encourage epithelial cell growth between the collagen layer and the silicone layer, then the silicone layer is spontaneously ejected soon after the wound is fully covered by neoepidermal skin. This eliminates the need for cutting, peeling, or any other delicate or invasive procedures to remove the silicone layer.
Of course, porous materials can be produced which are based upon other polymers~ including natural poly-mers such as cellulose and synthetic polymers which do not flow under elevated temperatures. Another protein which can be employed is leather, including ground leather obtained from leather scrap. A porous oam containing ground leather could be particularly useful as acoustic and/or thermal insulation~
Porosity can be achieved by any known technique, 2Q such as by forming a foam employing a blowing agent.
As mentioned above, freze-drying is often the preferred method for preparing foams of biocompatible materials.
This invention can be further and more specific-ally illustrated by the following example.
Bovine hide collagen in pulveri~ed form (0.55 g) was dispersed in 200 ml of 0.05M acetic in a refrigerated Eberbach blender over 1 hour. Chrondroitin 6-sulfate (o.044 g) was dissolved in 40 ml of 0.05M acetic acid and the solution was added dropwise to the stirred col-lagen dispersions. After 10 min of additional homo-yenization the blender was stopped and the dispersion ,, ~ 1 70~ 1 was poured into a 250 ml plaatic centrifuge bottle and was centrifuged for 1 hour at 2200 rpm at 4C in an International Model PR-l centrifuge. Following centrif-ugation, 140 ml were decanted from the bottle and the concentrate was blended for 15 min. 65 ml of the dispersion was poured into a shallow aluminum pan, 18x5 x 8.5 cm, which was placed on the shelf, maintained at -55C, of a Virtis 10 LN freeze drying chamber for 1 hour.
The frozen dispersion was then freeze dried to he point where no ice formed on the condenser coils of the chamber~ The dry foam was placed in a vacuum oven maintained at 105C, pressure 60 mtoor, for 12 hours. After allowing for 30 minutes of cooling the foam was covered with a 15-mil thick layer of room-temperature-curing silicone rubber medical adhesive sold under the trade mark SILASTIC by Dow Corning Corporation. It was then immediately placed in 0.05M
acetic acid at 4C for 48 hours to re-swell. The swollen membrane was then immersed in 200 ml of 0.25%
glutaraldehyde in 0.05M acetic acid for 24 hours at 20C, rinsed thoroughly with water and stored in 70/30 isopropanol/water until r ady for use as a closure for full-thickness skin wounds.
When prepared in this manner, the membrane exhib-its very high porosity when studied by a scanning elec-tron microscope. For example, pore volume fractions in the ran~e of 90-98% have been routinely determined by use of techniques well known to users of light and electron microscopes. The average value of pore diame-ter in these membranes has ranged from about 5 ~m to about 300 ~m.
r~3 ~ 1 7~0Q 1 In the absence of the heat-treatment step under vacuum following freeze drying, these membranes were observed to shrink, when immersed in an aqueous medium, le.g., 70/30 isopropanol/water) to about one-fourth or less of their original dimensions and to have irreversibly lost their porosity, as shown by scanning electron microscopy.
A total of approximately 11 membranes, pre-pared as described in Example 1 with variations to determine the effects of specific pararnetersJ were grafted onto skin wounds on female ~artley guinea piys.
The typical skin wounds were 1.5 x 3 cm, and were produced by removing the entixe epidermis and dermis to the panniculus carnosus. Each membrane was careflllly draped across the wound, sutured into place, and covered with a bandage. After varying per:iods of time, some of the animals were sacrificed and cross sectional slices of the graft were removed and studied. Histo-logical analyses indicated that collagen lattice porosity in excess of about 90% tends to encourage rapid cell growth and migration within the lattice, and that porosity of about 95% or higher tends to promote epithel~ial cell growth between the collagen lattice and the silicone layer. Other grafted wounds were allowed to heal completely. It was observed that epithelial cell growth between the collagen lattice and the silicone layer tends to cause the silicone layer to be spontaneously ejected soon after the neoepidermal layer closes the wound. This eliminates the need for peeling or surgically removing the silicone layer from the wound.
~ 3 ~00~ ~
Industrial Utility The porous protein foams produced by this inven-tion have utility in medical and surgical application requiring films, membranes, sutures, or other prosthe-ses which are biocompatible and porous. They are also believed to have utility as protein component in many foodstuffs. Other porous materials, such as non-woven materials, have use as fabrics, leather substitutes, etc.
1~ Equivalents Those skilled in the art will recognize, or be able tc ascertain employin~ no more than routine ex-perimentation, many equivalents to the specific materi-als, steps, etc., described above. Such equivalents are intended to be covered by the following claims.
Description MEI'HOD FOR PRESERVING POROSITY IN POROUS MATERIALS
Government Support The inventiorl described herein was made in the course of or under grants from the National Institutes of Health.
Field of_the Invention This invention is in the field of materials and more particularly relates to the treatment of porous materials to preserve their porosity.
Backqround of the invention Many materials are formed to contain a large number of poresO For example, certain materials or non-woven materials are porous, as are a large number of foams based upon natural or synthetic polymers. In many applications for such porous materials, it is important for the porous material to retain its porosity during exposure of the material to liquids without collapse of the pores.
One example of materials where this is true is a new class of tissue-compatible materials which are also insoluble in the presence of body fluids and controllably degradable in the presence of body enzymes and has been disclosed in U.S. Patent No. 4,280,954.
These materials are known as crosslinked collagen-mucopolysaccharide composites. They are synthe-sized by intimately contacting collagen with a muco-polysaccharide and subsequently crosslinking the resulting productO Suitable collagens can be derived .
1 1 ~0 ~
from a number of animal sources, and suitable mucopoly-saccharides include, but are not limited to, chondroi-tin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate, keratan sulfate, heparan sulfate, heparin, and hyalu ronic acid. Insolublization can be achieved by chemi-cal, radiation, or other suitable crosslinking tech-niques, or dehydrothermal treatment. Dehydrothermal treatment is particularly preferred and is achieved by reducing the moisture level of the composites to a very low level, such as by subjecting the composite material to elevated temperatures and high vacuum.
~ hese crosslinked collagen-mucopolysaccharide com posites are believed to be comprised of collagen mole-cules or collagen fibrils with long mucopolysaccharide chains attached to them. Crosslinking appears to an-chor the mucopolysaccharide chains to the collagen so they will not elute or otherwise become disengaged.
These crosslinked collagen-mucopolysaccharide com-posites have been found to retain the advantageous properties of native collagen. Additionally, such ma-terials can be synthesized to be very weakly antigenic, degradable by collagenase or other enzymes at controll-able rates, and insoluble in the presence of body fluids. Additionally, such composites can be synthe-sized to have ultimate tensile strengths, elongationsat break, and other mechanical properties particularly desired for artificial skin grafts and wound dressings.
1 ~J~
These crosslinked collagen-mucopolysaccharide compos;tes have been combined with moisture trans-mission control layers adherently bonded thereto to form synthetic skin. The moisture transmission con-trol layers are formed from nontoxic materials whichcontrol moisture flux of the overall membrane and can be formed from synthetic polymers such as silicone resins, polyacrylate or polymethacrylate esters or their copolymers, and polyurethanes. Such synthetic skin is described in U.S. Patent No. 4,060,081, issued to Yannas et al. on November 29, 19770 In many applications for the crosslinked collagen-mucopolysaccharide composites, it has been found highly preferable to prepare these materials as highly porous foams. For example, in the use of these materials as medical prostheses, including their use as synthetic skin, there is frequently a need for migration of cells from adjacent host tissue into tha prosthesis.
Such migration provides firmer attachment of the prostheses to the tissue and is also indispensable whenever there is a requirement for the invading cells to synthesize new functional tissue inside the pores of the prosthesis, which, if biodegradable, eventually disappears from the original site leaving in its place the newly synthesized tissue. In short, it has been found that high porosity in these composite materials, very oten in excess of 90% pore volume, allows a significantly greater degree of cell infiltration, elicits a much reduced fibrous sac, and allows desired tissue synthesis to occur at a much faster rate than corresponding material produced as membranes without high porosity.
., ,."~
~ ~ ~looo ~
Despite the need for porous materials, most tradi~
tional technqiues for producing foams are not suitable to produce materials which are biocompatible. For ex-ample, it is well known to produce foamed solids by em-ploying blowing agents to produce synthetic polymericfoams. An example of the use of blowing agents is the formation of polyurethane foams. However, the prepara-tion of such foamed polyurethanes generally involves the use of toxic chemicals, such as diisocyanates, which may often remain in an incompletely reacted form. This would be particularly objectionable in the case of a collagen or collagen-mucopolysaccharide based material, which would evoke little or no inflammatory response itself, but would generate a toxic inflamma-tory response if unreacted diisocyante were present.
It is also difficult to control foam density or poros-ity using blowing agents, and even when possible it re-quires elaborate processing steps including addition of catalytic systems which also may be toxic.
Because of the problems with conventional foam generation techniques, it has been found preferable to produce porous materials based upon crosslinked colla-gen mucopolysaccharide composites by a technique known as freeze drying. In this technique, an aqueous dis-persion of the composite is quickly frozen and the re-sulting ice particles are subsequently caused to sub-lime in the presence of vacuum. A solid, highly porous material results, and the degree of porosity can be controlled by adjusting the concentration of solids in the dispersion prior to the rapid freezing, as well as by adjusting the temperature and vacuum to which the drying membrane is exposed during the process.
Although the freeze drying technique has been found to be generally suitable, the materials produced do suffer one disadvantage. This disadvantage is that the dry porous solids obtained by sublimation of ice under vacuum often shrink considerably and irreversibly when brought into contact with liquids, such as an aqueous solution. Such shrinkage causes closure of the pores and makes the material less useful in the appli-cations where the high level of porosity is required or preferable. Unfortunately, most applications for these porous biocompatible materials require that the mate-rial be further processed or stored in aqueous solu-tions, or require that the materials be placed in con-tact with aqueous body fluids during use which would also cause undesirable shrinkage of a prosthesis formed from such materials.
The problems of shrinkage and pore collapse suf-fered by porous crosslinked collagen-mucopolysaccharide materials are illustrative of problems encountered with porous materials based upon a wide variety of natural and synthetic polymers when such porous materials are brought into contact with liquids.
Disclosure of the Invention This invention relates to the treatment of porous materials, particularly highly porous materials (e.g., above 90% pore volume) with a combirlation of elevated temperature and vacuum to modify such materials so that their porosity ls preserved when they are subsequently contacted with fluids.
, . . .
0 1~ ~
More particularly, the invention provides a method of producing a dimensionally-stable, non-collapsible highly porous foam based upon insoluble protein-based polymeric material, which comprises the steps of:
a~ forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated temperatures;
b) quickly freezing the dispersion to form frozen liquid particles;
c) subliming the frozen liquid particles to produce a highly porous foam;
d) subjecting the highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize the highly porous foam so th~t its porosity is pre-served when it is contacted with a liquid solution of a chemical cross-linking agent, and, e) subjecting the stabilized highly porous foam to a liquid solution of a chemical cross-linking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
An irnportant advantage is the degree of control over the porosity of the resulting protein foams which can be gained. Porosity, for example, can be con-trolled by adjustment of the solids conten~ of the dis~
persion prior to the quick freezing step as well as by adjustment of the temperature and pressure employed during the freeze-drying process.
Porosity is preserved, even upon contact with liquids, by the treatment under elevated temperature and vacuurn. This prevents shrinkage of the foam materials when they are subsequently conkacted with liquids during further processing, storage or use.
This is especially advantageous since the most cornmon methods of cross]inking collagen involve contacting it with an aqueous solution of a crosslinking agent, such as glutaraldehyde. Without prior dehydrothermal treatment, it would be very difficult to crosslink , the collagen without damaging its porosity. In addition, one preferred method of storing the collagen membrane described in U.S. Patent 4,060,081 (Yannas et al, 1977) is in the hydrated form, in a sterile watertight pouch that contains a piece of membrane immersed in a saline solution.
One class of materials suitable for production of porous protein foams is the class of crosslin~ed colla-gen-mucopolysaccharide composite materials described above. However, protein foams can be formed from pure collagen or other proteins in their pure form, such as gelatin, albumin, fibrinogen, and soybean protein~
Additionally, collagen or these other protein molecules can be grafted with various comonomers to form composite or grafted protein foams.
The initial step in preparation of these protein foams is the formation of an aqueous dispersion of the protein. Collagen based material can be employed by first swelling the collagen in aqueous acidic medium.
In this regard, it is particularly preferred to swell collagen at a low p~, such as about 3.5 or lower. See, examples 16 and 17 of U.S~ Patent No. 4,280,954.
Control over porosity of the foams can be achieved at the dispersion ~tage by adjusting the solid content of the dispersion~ ~n general, higher porosity will be attained as the solids content is lowered. Addition-ally, control of porosity can be obtained by adjusting the temperature and vacuum during freeze-drying.
In regard to producing shaped articles from the dispersions of protein, there are several alternative methods which mày be employed. In one method, the aqueous dispersion is simply filtered to produce porous sheets which can subse~uently be shaped. On the other hand, more intricate shapes, such as are often required to produce arterial or benous tubing, can be produced by a crossflow filtration molding method described in U.S. Patent No. 4,252,759.
. ' .
1 ~ ~00~ ~, Porosity is achieved by freeze drying techniques which generally involve subjecting the shaped article to low temperatures, so that ice crystals are formed, followed by sublimation of the ice under vacuum. Such techniques are known to those skilled in the art and many of these have been described in the patent litera-ture. See, for example, U. S. Patents 3,632,371;
3,471,598; 3r368,911; and 2,610,625.
The shaped protein foams are treated under high temperature and vacuum. In general, temperatures of 80C to 180C have been found suitable for collagen based foams. The vacuum may vary from about 1 mtorr up to slight vacuum just below atmospheric pressure.
Practical combinations of temperatures and pressures for collagen based materials have been found to be 80C
at 50 mtorr, 105C at 1 torr and 150C at 600 torr~ An increase in temperature or vacuum can be used singly or in combination to accelerate the process of pore pre-servationO
As mentioned above, a particularly important use for the highly porous protein foams produced by the method described herein i5 in multilayer membranes use-ful to cover wound dressing. Thus, wo~nd dressings or synthetic skin can be produced by multilayer membrane formed from either collagen or crosslinked collagen-mucopolysaccharide composite used in conjunction with moisture transmisslon control layers as described in U. S. 4,060,081.
When used as synthetic skin, the multilayer 3a membrane described in U. S. 4,060,081 exhibits two very beneficial characteristics that result from very high porosity. First, if the porosity of the collagen material exceeds about 90%, and is preferably about 95% t~ 98%, then epithelial cells tend to grow between the collagen layer and the silicone layer.
~l~oonl .
Since the collagen layex must be covered ~y epithelial cells before it can be biodegraded and the wound fully closed, this is very desirable. Second, if the porosity is sufficiently high to encourage epithelial cell growth between the collagen layer and the silicone layer, then the silicone layer is spontaneously ejected soon after the wound is fully covered by neoepidermal skin. This eliminates the need for cutting, peeling, or any other delicate or invasive procedures to remove the silicone layer.
Of course, porous materials can be produced which are based upon other polymers~ including natural poly-mers such as cellulose and synthetic polymers which do not flow under elevated temperatures. Another protein which can be employed is leather, including ground leather obtained from leather scrap. A porous oam containing ground leather could be particularly useful as acoustic and/or thermal insulation~
Porosity can be achieved by any known technique, 2Q such as by forming a foam employing a blowing agent.
As mentioned above, freze-drying is often the preferred method for preparing foams of biocompatible materials.
This invention can be further and more specific-ally illustrated by the following example.
Bovine hide collagen in pulveri~ed form (0.55 g) was dispersed in 200 ml of 0.05M acetic in a refrigerated Eberbach blender over 1 hour. Chrondroitin 6-sulfate (o.044 g) was dissolved in 40 ml of 0.05M acetic acid and the solution was added dropwise to the stirred col-lagen dispersions. After 10 min of additional homo-yenization the blender was stopped and the dispersion ,, ~ 1 70~ 1 was poured into a 250 ml plaatic centrifuge bottle and was centrifuged for 1 hour at 2200 rpm at 4C in an International Model PR-l centrifuge. Following centrif-ugation, 140 ml were decanted from the bottle and the concentrate was blended for 15 min. 65 ml of the dispersion was poured into a shallow aluminum pan, 18x5 x 8.5 cm, which was placed on the shelf, maintained at -55C, of a Virtis 10 LN freeze drying chamber for 1 hour.
The frozen dispersion was then freeze dried to he point where no ice formed on the condenser coils of the chamber~ The dry foam was placed in a vacuum oven maintained at 105C, pressure 60 mtoor, for 12 hours. After allowing for 30 minutes of cooling the foam was covered with a 15-mil thick layer of room-temperature-curing silicone rubber medical adhesive sold under the trade mark SILASTIC by Dow Corning Corporation. It was then immediately placed in 0.05M
acetic acid at 4C for 48 hours to re-swell. The swollen membrane was then immersed in 200 ml of 0.25%
glutaraldehyde in 0.05M acetic acid for 24 hours at 20C, rinsed thoroughly with water and stored in 70/30 isopropanol/water until r ady for use as a closure for full-thickness skin wounds.
When prepared in this manner, the membrane exhib-its very high porosity when studied by a scanning elec-tron microscope. For example, pore volume fractions in the ran~e of 90-98% have been routinely determined by use of techniques well known to users of light and electron microscopes. The average value of pore diame-ter in these membranes has ranged from about 5 ~m to about 300 ~m.
r~3 ~ 1 7~0Q 1 In the absence of the heat-treatment step under vacuum following freeze drying, these membranes were observed to shrink, when immersed in an aqueous medium, le.g., 70/30 isopropanol/water) to about one-fourth or less of their original dimensions and to have irreversibly lost their porosity, as shown by scanning electron microscopy.
A total of approximately 11 membranes, pre-pared as described in Example 1 with variations to determine the effects of specific pararnetersJ were grafted onto skin wounds on female ~artley guinea piys.
The typical skin wounds were 1.5 x 3 cm, and were produced by removing the entixe epidermis and dermis to the panniculus carnosus. Each membrane was careflllly draped across the wound, sutured into place, and covered with a bandage. After varying per:iods of time, some of the animals were sacrificed and cross sectional slices of the graft were removed and studied. Histo-logical analyses indicated that collagen lattice porosity in excess of about 90% tends to encourage rapid cell growth and migration within the lattice, and that porosity of about 95% or higher tends to promote epithel~ial cell growth between the collagen lattice and the silicone layer. Other grafted wounds were allowed to heal completely. It was observed that epithelial cell growth between the collagen lattice and the silicone layer tends to cause the silicone layer to be spontaneously ejected soon after the neoepidermal layer closes the wound. This eliminates the need for peeling or surgically removing the silicone layer from the wound.
~ 3 ~00~ ~
Industrial Utility The porous protein foams produced by this inven-tion have utility in medical and surgical application requiring films, membranes, sutures, or other prosthe-ses which are biocompatible and porous. They are also believed to have utility as protein component in many foodstuffs. Other porous materials, such as non-woven materials, have use as fabrics, leather substitutes, etc.
1~ Equivalents Those skilled in the art will recognize, or be able tc ascertain employin~ no more than routine ex-perimentation, many equivalents to the specific materi-als, steps, etc., described above. Such equivalents are intended to be covered by the following claims.
Claims (8)
1. A method of producing a dimensionally-stable, non-collapsible highly porous foam based upon an insol-uble protein-based polymeric material, comprising:
a. forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated temperatures;
b. quickly freezing said dispersion to form frozen liquid particles;
c. subliming said frozen liquid particles to produce a highly porous foam;
d. subjecting said highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize said highly porous foam so that its porosity is pre-served when it is contacted with a liquid solution of a chemical cross-linking agent; and, e. subjecting said stabilized highly porous foam to a liquid solution of a chemical cross-linking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
a. forming a liquid dispersion of an insoluble protein-based polymeric material which does not flow under elevated temperatures;
b. quickly freezing said dispersion to form frozen liquid particles;
c. subliming said frozen liquid particles to produce a highly porous foam;
d. subjecting said highly porous foam to elevated temperature and vacuum conditions sufficient to stabilize said highly porous foam so that its porosity is pre-served when it is contacted with a liquid solution of a chemical cross-linking agent; and, e. subjecting said stabilized highly porous foam to a liquid solution of a chemical cross-linking agent so as to produce a dimensionally-stable, non-collapsible highly porous foam material.
2, A method according to claim 1, wherein said liquid dispersion comprises an aqueous dispersion.
3. A method according to claim 2, wherein said insolubleprotein-based polymericmaterial comprises cross-linked collagen-mucopolysaccharide composite material.
4. A method according to claim 2, wherein said insoluble protein-based polymeric material comprises ground leather.
5. A method according to claim 4, wherein said highly porous foam has a porosity greater than about 95%.
6. A dimensionally-stable, non-collapsible highly porous foam material produced by 2 method as defined in claim 1.
7. A highly porous foam according to claim 6 having a porosity greater than about 95%.
8. In the production of a multi-layer membrane having a first layer formed from an insoluble protein-based polymeric material which does not flow under elevated temperatures and a second layer comprising a moisture transmission control layer, the improvement wherein said first layer comprises a highly porous foam material having a porosity greater than about 95%
which has been subjected to elevated temperature and vacuum in order to preserve its porosity and subse-quently contacted with an aqueous solution of a chemical crosslinking agent to cause said highly porous foam to become dimensionally-stable and non-collapsible.
which has been subjected to elevated temperature and vacuum in order to preserve its porosity and subse-quently contacted with an aqueous solution of a chemical crosslinking agent to cause said highly porous foam to become dimensionally-stable and non-collapsible.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US16989780A | 1980-07-17 | 1980-07-17 | |
| US169,897 | 1980-07-17 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1170001A true CA1170001A (en) | 1984-07-03 |
Family
ID=22617666
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000381746A Expired CA1170001A (en) | 1980-07-17 | 1981-07-15 | Method for preserving porosity in porous materials |
Country Status (1)
| Country | Link |
|---|---|
| CA (1) | CA1170001A (en) |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4861714A (en) * | 1985-04-04 | 1989-08-29 | Verax Corporation | Weighted collagen microsponge for immobilizing bioactive material |
| US4880429A (en) * | 1987-07-20 | 1989-11-14 | Stone Kevin R | Prosthetic meniscus |
| US5007934A (en) * | 1987-07-20 | 1991-04-16 | Regen Corporation | Prosthetic meniscus |
| EP0440198A2 (en) * | 1990-01-31 | 1991-08-07 | Gunze Limited | Process for preparation of dried collagen sponge |
| US5100783A (en) * | 1985-05-10 | 1992-03-31 | Verax Corporation | Weighted microsponge for immobilizing bioactive material |
| US5158574A (en) * | 1987-07-20 | 1992-10-27 | Regen Corporation | Prosthetic meniscus |
| US5263984A (en) * | 1987-07-20 | 1993-11-23 | Regen Biologics, Inc. | Prosthetic ligaments |
| US5306311A (en) * | 1987-07-20 | 1994-04-26 | Regen Corporation | Prosthetic articular cartilage |
| US5681353A (en) * | 1987-07-20 | 1997-10-28 | Regen Biologics, Inc. | Meniscal augmentation device |
| US5735902A (en) * | 1987-07-20 | 1998-04-07 | Regen Biologics, Inc. | Hand implant device |
-
1981
- 1981-07-15 CA CA000381746A patent/CA1170001A/en not_active Expired
Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4861714A (en) * | 1985-04-04 | 1989-08-29 | Verax Corporation | Weighted collagen microsponge for immobilizing bioactive material |
| US5100783A (en) * | 1985-05-10 | 1992-03-31 | Verax Corporation | Weighted microsponge for immobilizing bioactive material |
| US5624463A (en) * | 1987-07-20 | 1997-04-29 | Regen Biologics, Inc. | Prosthetic articular cartilage |
| US5007934A (en) * | 1987-07-20 | 1991-04-16 | Regen Corporation | Prosthetic meniscus |
| US5158574A (en) * | 1987-07-20 | 1992-10-27 | Regen Corporation | Prosthetic meniscus |
| US5263984A (en) * | 1987-07-20 | 1993-11-23 | Regen Biologics, Inc. | Prosthetic ligaments |
| US5306311A (en) * | 1987-07-20 | 1994-04-26 | Regen Corporation | Prosthetic articular cartilage |
| US4880429A (en) * | 1987-07-20 | 1989-11-14 | Stone Kevin R | Prosthetic meniscus |
| US5681353A (en) * | 1987-07-20 | 1997-10-28 | Regen Biologics, Inc. | Meniscal augmentation device |
| US5735902A (en) * | 1987-07-20 | 1998-04-07 | Regen Biologics, Inc. | Hand implant device |
| US5735903A (en) * | 1987-07-20 | 1998-04-07 | Li; Shu-Tung | Meniscal augmentation device |
| US6042610A (en) * | 1987-07-20 | 2000-03-28 | Regen Biologics, Inc. | Meniscal augmentation device |
| EP0440198A2 (en) * | 1990-01-31 | 1991-08-07 | Gunze Limited | Process for preparation of dried collagen sponge |
| EP0440198A3 (en) * | 1990-01-31 | 1992-01-15 | Gunze Limited | Process for preparation of dried collagen sponge |
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