CA1147507A - Composite material of de-n-acetylated chitin and fibrous collagen - Google Patents
Composite material of de-n-acetylated chitin and fibrous collagenInfo
- Publication number
- CA1147507A CA1147507A CA000373335A CA373335A CA1147507A CA 1147507 A CA1147507 A CA 1147507A CA 000373335 A CA000373335 A CA 000373335A CA 373335 A CA373335 A CA 373335A CA 1147507 A CA1147507 A CA 1147507A
- Authority
- CA
- Canada
- Prior art keywords
- collagen
- composite material
- aqueous
- acetylated chitin
- chitin
- 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.)
- Expired
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/10—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a carbohydrate
-
- A—HUMAN NECESSITIES
- A22—BUTCHERING; MEAT TREATMENT; PROCESSING POULTRY OR FISH
- A22C—PROCESSING MEAT, POULTRY, OR FISH
- A22C13/00—Sausage casings
- A22C13/0013—Chemical composition of synthetic sausage casings
-
- A—HUMAN NECESSITIES
- A22—BUTCHERING; MEAT TREATMENT; PROCESSING POULTRY OR FISH
- A22C—PROCESSING MEAT, POULTRY, OR FISH
- A22C13/00—Sausage casings
- A22C13/0013—Chemical composition of synthetic sausage casings
- A22C13/0016—Chemical composition of synthetic sausage casings based on proteins, e.g. collagen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/20—Polysaccharides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/26—Mixtures of macromolecular compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L33/00—Antithrombogenic treatment of surgical articles, e.g. sutures, catheters, prostheses, or of articles for the manipulation or conditioning of blood; Materials for such treatment
- A61L33/0005—Use of materials characterised by their function or physical properties
- A61L33/0011—Anticoagulant, e.g. heparin, platelet aggregation inhibitor, fibrinolytic agent, other than enzymes, attached to the substrate
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
- C08L5/00—Compositions of polysaccharides or of their derivatives not provided for in groups C08L1/00 or C08L3/00
- C08L5/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S128/00—Surgery
- Y10S128/08—Collagen
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S128/00—Surgery
- Y10S128/22—Blood coagulation
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S623/00—Prosthesis, i.e. artificial body members, parts thereof, or aids and accessories therefor
- Y10S623/915—Method or apparatus for preparing biological material
- Y10S623/916—Blood vessel
- Y10S623/917—Collagen
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
- Y10T428/00—Stock material or miscellaneous articles
- Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
- Y10T428/00—Stock material or miscellaneous articles
- Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
- Y10T428/2913—Rod, strand, filament or fiber
- Y10T428/2933—Coated or with bond, impregnation or core
- Y10T428/2964—Artificial fiber or filament
Abstract
ABSTRACT OF THE DISCLOSURE:
A composite material of de-N-acetylated chitin and fibrous collagen, which may be prepared by bringing the de-N-acetylated chitin and the fibrous collagen into mutual contact in an aqueous acidic medium followed by deacidifying the obtained product, the fibrous collagen being able to be partially replaced by gelatin and/or soluble collagen, and a shaped material derived from the composite material is excellent in mechanical strength, heat-resistance and biostability and advantageously employed in the field of medical materials, edibles such as edible casing and base materials for inmobilizing enzyme.
A composite material of de-N-acetylated chitin and fibrous collagen, which may be prepared by bringing the de-N-acetylated chitin and the fibrous collagen into mutual contact in an aqueous acidic medium followed by deacidifying the obtained product, the fibrous collagen being able to be partially replaced by gelatin and/or soluble collagen, and a shaped material derived from the composite material is excellent in mechanical strength, heat-resistance and biostability and advantageously employed in the field of medical materials, edibles such as edible casing and base materials for inmobilizing enzyme.
Description
11~75~
"
This invention relates to a novel composite material comprising de~N-acetylated chitin and fibrous collagen which ¦ may be partially replaced by gelatin or soluble collagen.
More particularly, the invention relates to a composite ~1 derived from the combination of the de-N-acetylated chitin, the fibrous collagen, the gelatin and the soluble collagen, which is excellent in mechanical strength a~d heat-resistance ¦and is not absorbed by a living body.
A representative protein of collagen is a scleroprotein lo contained in the connective tissues, bones, teeth, ligaments, tendons, cutises, fasciae, etc. of mammals, birds, etc. and broadly employed in the forms o edible casing material, threads for surgical suture, pieces for vasculoplastic trans-plantation or artificial skins, etc. Collagen is superior to other materials used in such fields, however, it is not always satisfactory for those purposes.
For instance, as a casing material for foods such as ham and sausage which necessitate smoke-drying~ gut of cattle, swine, sheep, etc., i.e., natural collagen material 1 has been utilized. However, there is a limit in the production I I of a shaped material from the natural collagen material and since the shaped material is produced via various complicated and troublesome steps of treatment, it is highly expensive.
Besides, since the shape and quality of natural material of collagen are irregular, respectively, such a material of natural ¦¦collagen has scarcely been applied on a conventional high-speed : :
11~7S~7 .
meat-stuffing machine in the production of ham and sausage.
A tubely shaped material made of collagen fibers, i.e., an artificial collagen casing has been proposed to resolve the above-mentioned demerits. However, such an arti-ficial collagen fiber is poor in film-farming property.
Besides, such an artificial collagen casing is large in thickness because of the necessity of retaining its mechanical strength Il and accordingly, there are demerits of not so good appearance ¦l and of remarkably impairing the dietary feeling. Further, in lo ¦¦ hams and sausages enclosed with such an artificial collagen ¦~ casing, unfavorable phenomena of breaking during its heating for cooking and of ablation of the casing from the stuffed meat take place.
Recently, a process for further subjecting the artifi-I cial collagen casing to a treatment of crosslinking has also been proposed for resolving the problem in cooking. However, such a crosslinked collagen casing is not satisfactory in the l dietal feeling, either.
- On the other hand, surgical materials such as the thread for surgical suture, artificial skin and hemostatic material, etc. from the collagen material are still unsatis-factory in the point of biostability (a property of not being absorbed by living body) other than the problem of strength.
The process for providing a biostability to a collagen material has been studied from various points, and there is a process having a step of crosslinking collagen. However, the shaped material from the thus cross-linked collagen is remarkably poorer in elasticity than those of conventional shaped material
"
This invention relates to a novel composite material comprising de~N-acetylated chitin and fibrous collagen which ¦ may be partially replaced by gelatin or soluble collagen.
More particularly, the invention relates to a composite ~1 derived from the combination of the de-N-acetylated chitin, the fibrous collagen, the gelatin and the soluble collagen, which is excellent in mechanical strength a~d heat-resistance ¦and is not absorbed by a living body.
A representative protein of collagen is a scleroprotein lo contained in the connective tissues, bones, teeth, ligaments, tendons, cutises, fasciae, etc. of mammals, birds, etc. and broadly employed in the forms o edible casing material, threads for surgical suture, pieces for vasculoplastic trans-plantation or artificial skins, etc. Collagen is superior to other materials used in such fields, however, it is not always satisfactory for those purposes.
For instance, as a casing material for foods such as ham and sausage which necessitate smoke-drying~ gut of cattle, swine, sheep, etc., i.e., natural collagen material 1 has been utilized. However, there is a limit in the production I I of a shaped material from the natural collagen material and since the shaped material is produced via various complicated and troublesome steps of treatment, it is highly expensive.
Besides, since the shape and quality of natural material of collagen are irregular, respectively, such a material of natural ¦¦collagen has scarcely been applied on a conventional high-speed : :
11~7S~7 .
meat-stuffing machine in the production of ham and sausage.
A tubely shaped material made of collagen fibers, i.e., an artificial collagen casing has been proposed to resolve the above-mentioned demerits. However, such an arti-ficial collagen fiber is poor in film-farming property.
Besides, such an artificial collagen casing is large in thickness because of the necessity of retaining its mechanical strength Il and accordingly, there are demerits of not so good appearance ¦l and of remarkably impairing the dietary feeling. Further, in lo ¦¦ hams and sausages enclosed with such an artificial collagen ¦~ casing, unfavorable phenomena of breaking during its heating for cooking and of ablation of the casing from the stuffed meat take place.
Recently, a process for further subjecting the artifi-I cial collagen casing to a treatment of crosslinking has also been proposed for resolving the problem in cooking. However, such a crosslinked collagen casing is not satisfactory in the l dietal feeling, either.
- On the other hand, surgical materials such as the thread for surgical suture, artificial skin and hemostatic material, etc. from the collagen material are still unsatis-factory in the point of biostability (a property of not being absorbed by living body) other than the problem of strength.
The process for providing a biostability to a collagen material has been studied from various points, and there is a process having a step of crosslinking collagen. However, the shaped material from the thus cross-linked collagen is remarkably poorer in elasticity than those of conventional shaped material
- 2 -.
~1~7507 of collagen. In addition, the following process is under ~tria~ with an intension of resolving the above-mentioned problems by forming the polyion complex between collagen and ¦la high polymeric substance having carboxyl group(s) or sulfa~e ¦Igroup(s):
I (~
H3N - ~ - C00 + Y - COo or Y - S03 ~ Y - C00 H3N - ~ - C00 or (~
Y - S03 H3N - ~ - C00 wherein H3N-~-C00 represents ion of collagen molecule and Y-C00 lo or Y-S03 represents an ion of high polymeric substance.
The thus obtained polyion complex is excellent in biostability and has an antithrombogenic property.~ However, it dissolves into an aqueous medium, for example, an aqueous solution of an inorganic salt such as sodium chloride and is poor in mechanical properties. Accordingly, it is necessary to subject the polyion complex further to a treatment for stabilization such as crosslinking, etc.
The inventors of the present invention, during the course of studying the development of new composite materials suitable for use in broad range of purposes while reviewing the actual state of the art, have found out that a two-component composite comprising de-N-acetylated chitin and fibrous collagen or a multi-conponent composite comprising de-N-acetylated chitin,
~1~7507 of collagen. In addition, the following process is under ~tria~ with an intension of resolving the above-mentioned problems by forming the polyion complex between collagen and ¦la high polymeric substance having carboxyl group(s) or sulfa~e ¦Igroup(s):
I (~
H3N - ~ - C00 + Y - COo or Y - S03 ~ Y - C00 H3N - ~ - C00 or (~
Y - S03 H3N - ~ - C00 wherein H3N-~-C00 represents ion of collagen molecule and Y-C00 lo or Y-S03 represents an ion of high polymeric substance.
The thus obtained polyion complex is excellent in biostability and has an antithrombogenic property.~ However, it dissolves into an aqueous medium, for example, an aqueous solution of an inorganic salt such as sodium chloride and is poor in mechanical properties. Accordingly, it is necessary to subject the polyion complex further to a treatment for stabilization such as crosslinking, etc.
The inventors of the present invention, during the course of studying the development of new composite materials suitable for use in broad range of purposes while reviewing the actual state of the art, have found out that a two-component composite comprising de-N-acetylated chitin and fibrous collagen or a multi-conponent composite comprising de-N-acetylated chitin,
- 3 -'' ~
:' 11~7S~)~
., fibrous collagen and gelatin and/or soluble collagen, which may be prepared by bringing each proteinous substance into contact with the de-N-acetylated chitin, is extremely suitable to resolve the problems in the prior art.
Accordingly, the present invention provides a two-component composite comprising de-~-acetylated chitin and fibrous llcollagen wherein the amount of the de-N-acetylated chitin and ¦¦the fibrous collagen is selected from the range of 0.01 to 99 and 1 to 99.99 parts by weight, respectively, to 100 parts lo by weight of the composite material, and provides a three- or four-component composite obtained by partially replacing the : fibrous collagen of the two-component composite material with gelatin and/or soluble collagen wherein the amount of the gelatin and/or soluble collagen is up to 40 parts by weight to 100 parts by weight of the composite material and the amount . ¦ of the de-N-acetylated chitin and the fibrous collagen is the I same as in the two component composite material. In addition, the present invention provides a method for preparing the above-mentioned composite material wherein the de-N-acetylated chitin and the proteinaceous substance of the fibrous collagen, the gelatin and the soluble collagen are brought into mutual contact in an aqueous acidic medium of pH 1 to 6, preferably : pH 3 to 6 and then the obtained product is deacidified followed by cross-linking the deacidified product, if necessary.
Hitherto, proteinic composite materials such as those comprising collagen fiber and a soluble protein or comprising .' ' ' 11475~)7 collagen fiber and gelatin, etc. have been reported. However, ~I since in such a proteinic composition, the bonding between collagen and the soluble component is weak, the reduction of ~ strength of such a composition is remarkable when the composition ¦¦ is brought into hot water. On the other hand, the composite material according to the present invention does not dissolve ¦ into an aqueous salt solution, and is excellent in mechanical ¦ strength which is not reduced particularly in hot water, in non-absorbability by living bodies, in blood-coagulating lo property, in resistance against the attack of bacteria, in film-forming ability and in close adhesion to stuffed materials such as meat.
Although it has not yet been elucidated why the com-posite material according to the present invention has an excellent mechanical properties, non-abso~bability~by living bodies and a blood-coagulating property, the existence of the strong bonding via the de-N-acetylated chitin and the proteinaceous substance of the fibrous collagen, the gelatin and the soluble collagen is presumed to form a stable polyion complex as follows:
:
., ~ (3 t3 OE) (3 ~
- H3N - ~ - COO + Z-NH3 > H3N - ~ - COO H3N - Z
~:: ~9 (3 I wherein H3N-~-COO represents an ion of the proteinaceous molecule and H3N-Z represents an ion of the de-N-acety-lated chitin molecule.
., _ 5 _ `
' 11475~)7 The composite material according to the present inven-l, tion is applicable to various usages ~s one of such usages, jl an edible casing material is mentioned. Besides, as Kiriyama ~¦ et al have reported in A.N.I. No. 71, since chitosan has the ¦, following physiological activities:
Z! a) returning the raised level of serum cholesterol to normal state, b) inhibiting the raising of the level of blood sugar, c) removing growth-inhibiting substances and lo d) preventing the colonal cancer, the composite material according to the present invention may be added to foods in the form of a spherically shaped material or a fibrous material, or may exhibit the preventive or therapeutic effects to the various symptoms such as a) to d) Il as an edible material.
I Furthermore, since the composite material according ¦¦ to the present invention is not absorbable by living bodies owing to one of its components, de-N-acetylated chitin, and also has a blood-coagulating property, it can be surgically or therapeutically applied in the forms of film, fiber and those having a three dimensional structure. Although the crosslinked or not-crosslinked composite material of the present invention is itself thrombogenic, it can be applied as an anti-thrombogenic material after being heparin-treated and ~¦ possibly applied as a medical material for artificial vessels, artificial skins, artificial kidneys, etc.
-' ~ ' '' ' .
:, 11~75V7 1, , In addition, since the composite material according to the present invention is excellent in adsorbing ~roteins, ,it is possibly applicable as a base material for inmobilizing ~lenzymes and microorganisms or as an adsorbent of enzymes and ¦Imicroorganisms.
The following are the more precise description of the present invention.
The de-N-acetylated chitin mentioned in the present invention is, from the viewpoint of solubility and processability, of a degree of de-N-acetylation of 50 to 100 ~, preferably 70 to 100 % and of a viscosity of 20 to 1000 cp, the viscosity being determined at 20-C on an aqueous 1 ~ solution of acetic acid containing O.S ~ by weight of the de-N-acetylated chitin.
Co~mercialized chitosan may be used or the product obtained ¦by treating chitin in an aqueous alkali solution of~ a high concentration while heating may be used, the chitin being obtained from shells of arthropods such as crabs, lobsters, - shrimps, etc. by a conventional manner of separation and I purification.
Protein mentioned in the present invention means two ` kinds of proteins, one of which i5 the fibrous collagen in-soluble in an aqueous solvent and the other is the protein ; soluble in the aqueous solvent, i.e., gelatin and soluble collagen. They are hereinafter referred to as the proteinic component. The proteinic component is obtainable by separating ¦and purifying protelnaceous tissues from living bodies of !
11475~)7 varlous animals, or by chemically treating the separated and purified protein. The aqueous solvent mentioned in the present invention is an aqueous solution of acid or alkali. As the acid, an inorganic acid such as hydrochloric acid, etc., a single organic acid such as acetic acid, propionic acid, adipic ~ acid or a mixture of more than two kinds of the organic acid ¦Ican be exemplified. The alkali means a conventional alkali ¦¦such as sodium hydroxide, potassium hydroxide, etc.
1~ Collagen fiber has been obtained by removing non-lo collagenic components from the fibrous components consti-tuting major protein which is present in the connecting tissues of animals with chemical and mechanical treatments.
Collagen fiber, for example, may be prepared by a process wherein bovine hide or bovine Achilles tendon, after having subjected to depilating teatment, is finely cut by a mincing machine and is swollen in an aqueous acidic or alkaline medium, and then is disintegrated by a grinder to obtain ~ an aqueous dispersion, followed by filtration, if neces-I sary.
In addition, it is preferable for the improvement of strength and homogenization of the produced composite material to subject the thus finely cut hide or tendon to a pretreatment.
As a process of the pretreatment, crosslinking of amino group of lysine residue in collagen by a crosslinking agent such as formaldehyde, glutaraldehyde, dialdehyde starch, glyoxal, smoke-drying liquid, epihalohydrin, etc., succinylation . .1 . ~ ' .
.
of the amino group by succinic anhydride, acylation of the amino group by carboxylic acid anhydride or esterification of carboxyl group in the side chain of aspartic acid residue ¦lor glutamic acid residue in collagen are mentioned.
Besides, as the raw material for collagen fiber, llchips and rubbishes remaining after preparation of natural ¦¦or artificial collagen casing, which are not utllizable for ¦the casing are mentioned.
The fibrous collagen in the present invention is of 1 to 3 ~ in diameter and of 0.1 to 15 mm in length and of 5,000 to 1,000,000, preferably 20,000 to 500,000 in molecular weight, however, not necessarily being restricted wlthin such I ranges.
On the other hand, the other proteinic component of gelatin in the present invention may be one used in an indus-trial scale in food industries. The soluble collagen herein-mentioned is soluble in an aqueous solvent and may be obtained by treating the fibrous collagen with proteinase, an acid or an alkali to be wholly soluble into the aqueous solvent.
2~ Besides, a partly solubilized product obtainable in the treat-ment of the fibrous collagen is usable as a mixture of the fibrous collagen and the soluble collagen.
'I'he soluble protein of the gelatin and the soluble collagen amounts to 0 to 40 parts by weight to 100 parts by weight of the composite material according to the invention.
When the amount of the gelatin, the soluble collagen or the ., !l _ g _ .
:
I :~1475~7 mixture thereof is more than 40 parts by weight, the moisture content of the obtalned composite material is so much that the 'maintaining of the shape of the composite material is difficult and the tensile strength is poor. The gelatin is preferable as the soluble protein, and the amount of the gelatin is preferably 5 to 30 parts by weight to 100 parts by weight of the composite l! material.
¦ The soluble protein, as a component of the composite l material, contributes to the pliability, the reduced thermal lo deformation temperature, the improved mechanical strength, and also the improved dietary feeling of the composite material.
The composite material according to the present inven-l tion is also available by at first bringing the de-N-acetylated - ~ chitin and the proteinic component into contact in a medium having pH value in the range of 1 to 6, preferably,of 3 to 6 and then deacidifying of the product thus obtained followed by subjecting to crosslinking, if necessary.
The deacidifying treatment mentioned herein means the ~ adjusting of the pH of the reaction product of the de-N-acetylated ; 20 chitin and the proteinic component higher than the isoelectric point of the reaction product. The deacidificating treatment is, for instance, an addition of an alkaline solution such as sodium hydroxide, potassium hydroxide or ammonium hydroxide to bring the pH higher than 7, an addition of an aqueous solution of alkaline salt, a removal of the acid present in the reaction ~mixture by ev poration or a treatment by electrodeposition.
, .
,.
~, . . .
, .
'' ~ , , .
.
li475{)7 The gelatin obtained by denaturing collagen has various isoelectric point, its jelly strength, its ash content, etc. according to the process of denaturation treatment.
~Accordingly, in the case where the proteinaceous composite ~Imaterial is prepared by an electrodeposition method from the ¦~gelatin obtained by an acid-treatment, it is necessary to adjust the pH of the electrodeposition-controlling liquid higher than 7 to make the shaped composite material on the cathode. On the other hand, in the case where the gelatin I lo obtained by alkali-treatment is used, the shaped composite material must be formed on the anode by adjusting the pH of the electrodeposition-controlling liquid lower than 7.
Whereas, in the present invention, the composite material can also be prepared by electrodeposition without the above-mentioned restriction owing to the action of the de-N-acetylated chitin which is one of the components of the composition. Meanwhile, as the gelatin for use in the case ~1~ of shaping by electrodeposition, a product containing less than 0.5 % by weight of ash, preferably less than 0.2 % by weight of ash is desirable from the viewpoints of quality and demand of electricity in preparation.
Besides, according to the process of the present I invention, by combining the proteinic component which has been insufficient in shape-forming property with the de-N-acetylated chitin, the shape-forming property, for example, membrane-forming property can be improved and the ll I
', - 11 -.
.
~1475V7 problems concerning the preparation of the composite material such as the yield in the preparation, the stability and the uniformity of the product are possibly improved. ~esides, I since the remnant free aldehyde after the pretreatment of ¦¦the fibrous collagen by the cross-linking is also reacted with ¦ the de-N-acetylated chitin according to the invention, the I composite material is desirable from the viewpoint of safety.
The following are the more detailed explanation in preparing the composite material of the present invention.
lo As one of the embodiments of the contact of the pro-teinic component and the de-N-acetylated chitin, a process comprising the steps of mixing an acid medium containing the de-N-acetylated chitin and a medium containing a proteinic component and dispersing the mixture can be exemplified. In this case, the concentration of the de-N-acetylated chitin in the acidic medium is less than 5 % by weight, preferably less than 1 % by weight in connection to its viscosity (the medium being an aqueous dilute solution of acetic acid, hydro-chloric acid, etc.), and the pH of the acidic medium is 1 to 6, preferably 3 to 6. It is not always necessary that the de-N-acetylated chitin is homogeneously dissolved in the medium, and the de-N-acetylated chitin may be dispersed uni~ormly as a fibrous state. The medium containing the proteinic com-ponent may be prepared by dispersing the proteinic component into an aqueous acidic solution of hydrochloric acid, etc.
and by adjusting the pH thereof at 3 to 6, preferably ,1 , -, 11~7507 3 to 4. However, the acidity of the medi~m is not necessarily required, that is, the medium may be neutral or alkaline. In Il this case, it is finally necessary that the p~ of the mixture ¦¦of the medium containing the de-N-acetylated chitin and the ¦medium containing the proteinic component is in the acidic ¦jregion. In addition, in the case of electrodeposition, the medium containing the proteinic component is used in an acidic ~ state.
¦ In the present invention, a film-like composite material lo is available by pouring the de-bubbled mixture of the medium containing the de-N-acetylated chitin and the medium containing the proteinic component onto a glass plate and then de-acidifying the poured mixture by the use of a drier with hot air. The film-like composite material is characterized in that its strength does not depend on the tensile direction, that is, longitudinal or transversal in spite of having the fibrous protein.
The above-mentioned characteristic feature of the film-~- like composite material according to the present invention 2~ can be exhibited by the presence of the de-N-acetylated chitin in the composite material.
I The spherically shaped composite material is prepared by dropping the de-bubbled mixture of the two media into a hydrophobic organic solvent such as toluene or xylene ` and then treating the thus ~ormed material with an aqueous alcoholic solution to deacidify the spherically shaped material.
,,1 '~' , - 13 -,.~
', 1~75~7 , .
Furthermore, fiber-like, film-like or hollowed composite material is possibly prepared by extruding the mixture fxom a hole-liXe or slit-like nozzle(s) into an aqueous solution of sodium chloride or ammonium hydroxide at a high concentration. In addition, the formed composite material of the present invention can be obtained by electrodeposition in which an aqueous medium containing the de-N-acetylated chitin and the proteinic com-ponent is introduced into an electrolytic cell provided with at least one cathode and at least one anode, and a direct voltage is loaded between both the electrodes to make the composite material accumulated on the surface of the predetermined lelectrode.
The tube-like composite material according to the present invention has an extremely thin wall rnembrane excellent in strength, and accordingly, it is highly suitable for edible casing material with a desirable dietary feeling.
In the case where the composite material according to the present invention is used for the edible casing, it is preferable that the amount of the de-N-acetylated chitin is selected from the range of 0.01 to 60 parts by weight, that of the fibrous collagen is selected from the range of 40 to 99.99 parts by weight and, if employed, that of the soluble protein of the gelatin, the soluble collagen or the mixture thereof is selected from the range of 5 to 30 parts by weight to 100 parts by weight of the composite material from the view points of the dietary feeling, the strength at smoke-drying ,,~
.
: ~ .
. ,, - 14 -.
~1~75~)7 1, and cooking, the thermal deformation temperature and the heat-stability of the casing. On the other hand, in the case where the composite material according to the present invention is ~used as a food additive, the amount of the de-N-acetylated chitin is selected from the range of 1 to 99 parts by weight to 100 parts by weight of the composite material and it is preferable to increase the amount of the de-N-acetylated chitin.
As another embodiment of the contact of the proteinic component and de-N-acetylated chitin, a process is exemplified in which a preliminarily shaped proteinic material into sheets, films, tubes, fiber-like forms, etc. is immersed into a acidified medium containing the de-N-acetylated chitin and then the shaped material is subjected to deacidifying treatment. In this case, ¦ since the superficial part of the composite materia~l is composed ¦ of the de-N-acetylated chitin c~mponent, such a shaped composite material shows extremely unique properties such as improved I mechanical properties, improved thermal resistance and anti-bacterial property.
Furthermore, in the case where the composite material according to the present invention obtained by one of the above-mentioned several methods is further subjected to a crosslinking treatment, the thus crosslinked composite material exhibits an improved property in blood-coagulation and an improved resistance to acids.
In the crosslinking treatment, a composite material : 11 . l ,' .
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il47507 according to the present invention is immersed into an a~ueous solution prepared by dissolving a crosslinking agent into a buffer solution of phosphoric acid at a pH of 7.4 for 0.1 to ll hour at 10 to 50-C. After the crosslinking is over, by ¦washing the crosslinked material with water, a water-insoluble ~crosslinked composite material is obtained.
Since it is possible by such a process of crosslinking to cause not only the crosslinking of the de-N-acetylated chitin but also the crosslinking between the de-N-acetylated chitin lo and the proteinic component, an improved bonding force in the surface of the composite material is obtained. The crosslinking occurs between the same or the different functional groups among amino groups and hydroxyl groups in the de-N-acetylated chitin and collagen. As a crosslinking agent, the same agents mentioned in the crosslinking carried out in the pretreatment ¦of the fibrous collagen are equally mentioned.
The present invention provides a novel composite material by combining the proteinic component and the de-N-`; acetylated chitin, which has excellent specific properties more than compensating the demerits of the de-N-acetylated chitin and the proteinic component, and the present invention contributes greatly to industries because the de-N-acetylated chitin is present in nature in abundance, however, owing to its high crystalinity and chemical stability, it has been utilized only in narrowly restricted fields.
,;
:' l ., , , ' - ' ' ~75()7 , ~I EXAMPLE 1 ' After immersing 100 kg of a salted steer hide into water for 15 hours, washing by water, and defatting, 90 kg of the thus treated hide was immersed into 450 kg of an aqueous liquid containing 2 ~ by weight of calcium hydroxide, 0.5 %
by weight of sodium sulfide and 0.5 % by weight of diethylamine Il at 25-C for 24 hours while gently stirring the mixture. The ¦ thus treated hide was subjected successively to a depilating I roll to remove hairs and decomposition products formed by the I treatment with calcium hydroxide, to a splitting machine to ~¦ roughly divide the hide, to a meat slicer to cut the divided I hide into tapes of 5 mm in width and then to a mincing machine to cut the tapes into 5 cm in length.
Eighty kilograms of the thus obtained hide (referred to the refined hide hereinafter) were dispersed in~o 800 kg of water, and 3 kg of acetic acid were added to the dispersion and then after stirring gently for 12 hours, the refined hide was separated from the liquid, dehydrated by centrifugation and washed with de-ionized water until the electric conductivity of the washings became less than 20~u/cm. At this time, the . ash content of the refined hide was less than 0.1 %. Forty kilograms of the thus treated refined hide were dispersed into 400 kg of an aqueous solution containing 0.015 % by weight of glutaraldehyde and adjusted to pH of 3.0 by the addition of hydrochloric acid, and then crosslinked while stirring the solution gently for 12 hours at 20C.
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11~L7507 ~~ The crosslinked refined hide was collected by filtra-tion with a wire netting and subjected to a pulp-refiner together with 113 kg of cold water to beat the fiber-bundles of the refined hide resulting in a dispersion of collagen fibers, f to which hydrochloric acid was further added to make the pH
tllereof 3Ø The thus obtained dispersion weighing 125 kg contained 2.5 % by weight of crosslinked collagen comprising a mixture of collagen fibers of 1 to 3 ~ in diameter and of 1 to 10 mm in length and still finer collagen fibriles.
lo Meanwhile, into 10 kg of an aqueous hydrochloric acid solution of pH of 3, one kg of de-N-acetylated chitin having a degree of de-N-acetylation of 95 % were dissolved and after adjusting the pH of the solution to 3.0, 20 kg of an aqueous solution of de-N-acetylated chitin were obtained.
Into 1.5 kg of the aqueous dispersion of crosslinked collagen fiber, 50 kg of de-ionized water were added and the mixture was stirred at a high speed, and then, 374 g or the : aqueous solution of de-N-acetylated chitin was added to the :~ dispersion and the mixture was stirred fGr one hour to be an ~. 20 aqueous mixture containing collagen fiber and de-N-acetylated ~:
` chitin at a weight ratio of 100/5, the viscosity and the . electric conductivity of the aqueous mixture being 200 cP
and 250 ~U /cm, respectively.
~:~ From the aqueous mixture of collagen fiber and de-N-I acetylated chitin, a tube-form membrane was formed by electro-,j - 18 -. ..: . . . . ,, , - - ,, . ~ , .
~75~)7 deposition in which a membrane was formed on the cathode (17.5 mm~) by electrophoresis of collagen fibex and de-N-acetylated chitin.
; The thus formed membrane was pulled up, at the velocity of pulling up being 10 m/min.
The thus prepared tube-form membrane just after elec-trodeposition contained 2 g of water per g of the dry membrane and showed a tensile strength of 200kg/cm2.
~¦ The pulled-up membrane, after immersing into an aqueous 7 ~ by weight of glycerol solution, was dried by a hot air at lo ¦l 75-C for 2 min, while holding the tube-form membrane by pressured air of 300 mm H2O, to be the composite material (Specimen A) - !l according to the present invention.
On the other hand, for comparison, a comparative ¦ tube-form membrane (Comparative Specimen A) consisting only of collagen fiber was prepared by the similar process mentioned above except for not adding de-N-acetylated chitin to crosslinked l collagen fiber. The Comparative Specimen A contained 7 g of ¦ water per g of the dry membrane just after electrodeposition showing a tensile strength of 100 kg/cm2.
The results of determination of tensile strength and tear strength in wet state of Specimen A and Comparative Specimen A according to the method described in JIS P-8113 and JIS P-8116 are shown in Table 1. As will be seen in Table 1, the specimen prepared from a mixture of crosslinked collagen fiber and de-N-acetylated chitin showed a remarkably improved ~ I tensile strength, and it was possible to obtain an extremely : li , ~ , .
.
~1~75V7 thin membrane.
Table 2 shows the results of strength-determination of the membranes by the same method as mentioned above after immersing into an aqueous 0.2N sodium chloride solution at 20C
for 24 hours. As will be seen in Table 2, the composite material according to the present invention is superior to the Comparative Specimen A in strength, which suggests not only the ionic bonding between collagen fiber and de-N-acetylated chitin but also another bonding factor.
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. Table 1 ,~
Specimen Thickness Tensile Tear strength Elongation at of membrane strength (g cm/cm) ~reak (11 ) (kg/cm2) l l . ~
Specimen A 10 460 36 32 l~ Compara-ll tivè 15 250 25 25 Specimen A
. . _ _ _ ~ Table_2 . . -- - -------- -----Specimen Tensile Tear strength Elongation at strength (g cm/cm) break (kg/cm2 ) ` ( ~ ) ._ Specimen A 275 61 115 Comparative 130 48 64 ',:
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~1475V7 I EXAMPLE 2:
. . _ One kg of an acid-treated gelatin (lot number of F-795, manufactured by Miyagi Chem. Co. Ltd., Japan) was dissolved into 10 kg of an aqueous hydrochloric acid solution I ll of pH of 3, and after adjusting the pH to 3.0, 20 kg of an Il aqueous solution of gelatin was obtained.
¦ After adding 50 kg of de-ionized water to 15 kg of ¦¦ the aqueous dispersion of c?rosslinked collagen fiber prepared ¦ in Example 1 and stirring rapidly, 374 g of the aqueous lo solution of de-N-acetylated chitin prepared in Example 1 and 374 g of the aqueous solution of gelatin were further added to the dispersion of crosslinked collagen fiber, and the whole ¦ mixture was stirred for one hour to obtain an aqueous mixture of collagen fiber, de-N-acetylated chitin and gelatin at a weight ratio of collagen: de-N-acetylated chitin:, gelatin of 100 : 5 : 5.
; From the thus obtained aqueous mixture, a tube-form membrane was formed by the same method as in Example 1 (Specimen B).
For comparison, on the other hand, a comparative tube-form membrane (Comparative Specimen B) consisting of collagen and gelatin was prepared by the same process, however, without adding de-N-acetylated chitin.
The results of determination of tensile strength and tear strength of the products by the same methods described ¦in Example 1 in wet state are shown in Table 3. As is seen in ,:; ,1 , 11~7507 Il Table 3, the strength of the membrane containing collagen 1, fiber, de-N-acetylated chitin and gelatin was improved as Il compared to that of the membrane only containing collagen ¦! fiber and gelatin, thus showing the effect of added de-N-acetylated chitin.
Furthermore, the results of determination of the mechanical properties of the products after immersing them in warm water at 40-C for 24 hours shown in Table 4 indi-cate the superiority of the composite material according to the present invention in strength to the product obtained without adding de-N-acetylated chitin.
.
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. ' , Table 3 Specimen Thickness Tensile Tear strength Elongation (~) strength (g cm/cm)at break (kg/cm2) ( %) , .. _ . .. _ .
Specimen B 10 480 36 35 Specimen B 10 340 27 27 Table 4 ... . _ _ Specimen Tensile strength Tear strength .Elongation at (kg/cm2) (g cm/cm) break (~) _ Specimen B 370 60110 : Specimen B 140 4560 `.~ I
' 11~75V7 EXAMPLE 3:
A tube-form composite material (Specimen C) was pre-pared in the same manner as in Example 1 except for the weight ¦ratio of the collagen fiber to de-N-acetylated chitin of 100/20 'l and a membrane-pulling-up velocity of 25 m/min. For comparison, a sigle material (a dispersion of collagen fiber of concentra-tion of 0.58 % by weight) was processed to be a membrane, however, the membrane could not be continuously pulled up.
The mechanical properties of Specimen C were: 8 ~ in thickness, 510 kg/cm2 in tensile strength, 32 % in elongation at break and 32 g cm/cm of tear strength.
EXAMPLE 4:
A tube-form composite material (Specimen D) was prepared in the same manner as in Example 1 and 2 except for the weight ratio of collagen fiber: de-N-acetylated chitin: gelatin of 100 : 20 : 5 and a membrane-pulling-up velocity of 25 m/min.
For comparison, an aqueous mixture of dispersed collagen fiber ; and gelatin of the weight ratio of collagen fiber: gelatin of 100 : 5 was tried to be a membrane, however, the membrane could not be continuously pulled up. The mechanical properties of S~ecimen D were: 8 ~ in thickness, 490 kg/cm2, in tensile strength, 33 % in elongation at break and 33 g cm/cm in tear strength.
EXAMPLE 5:
A tube-form composite material (Specimen E) was prepared in the same manner as in Example 1 and 2 except for using . ' , ' , : ' .. ~
~ 1475~)7 llalkali-treated gelatin (lot number of F-800, Miyagi Chem. Co.
Ltd., Japan) instead of acid-treated gelatin of Example 2 and the weight ratio of collagen fiber: gelatin: de-N-acetylated l chitin of 100 : 20 : 10 and a membrane-pulling-up velocity of 15 m/min. For comparison, an aqueous mixture of collagen ~I fiber and gelatin at a weight ratio of collagen to gelatin of ¦1 100 : 20 was subjected to electrodeposition, however, no membrane deposited on the electrode. The mechanical properties of Specimen E were: 9 ~ in thickness, 440 kg/cm2 in tensile strength, 34 % in elongation at break and 30 g cm/cm in tear strength.
EXAMPLE 6:
After mixing 10 Xg of the dispersion of crosslinked collagen fiber obtained in Example 1 and 1.5 kg of an aqueous ¦ 5 % by weight of de-N-acetylated chitin solution ob,tained in Example 1 while stirring, the mixture was de-bubbled under a reduced pressure. The de-bubbled mixture was extruded from a slit of lO cm in width and of 0.3 mm in height into an aqueous O.lN ammonium hydroxide solution contained in a coagulation bath. The thus obtained film-form material was washed with water, and was treated in the same manner as in Example 1 to be a film-form composite material according to the present invention (Specimen F).
The results of the determination in a manner as in Example 1 were: 30 ~ in thickness, 520 kg/cm2 in tensile stren~th, 30 % in elongation at break and 75 g cm/cm of tear strength.
`::
~1~75{)7 On the other hand, a film prepared from the same , dispersion of crosslinked collagen fiber in the same manner as above (Comparative Specimen C) without adding the solution of de-N-acetylated chitin showed the following properties:
30 ~ in thickness, 200 kg/cm2 in tensile strength, 26 % in elongation at break and 48 g cm/cm in tear strength.
From the above-mentioned results, the composite material ¦ according to the present invention was confirmed to be superior to the single material (only consisting of crosslinked collagen fiber) in mechanical strength.
EXAMPLE 7:
; After mixing 10 kg of the dispersion of crosslinked collagen fiber obtained in Example 1, 1 kg of an aqueous 5 %
by weight gelatin solution obtained in Example 2 and 1.5 kg ;~ of an aqueous 5 % by weight de-N-acetylated chitin solution obtained in Example 1, the mixture was well stirred and de-bubbled under a reduced pressure, and subjected to the same ¦ procedures as in Example 6 to obtain a film-form composite material according to the present invention (Specimen G).
The mechanical properties of Specimen G in wet state were:
30 ~ in thickness, 495 kg/cm in tensile strength, 31 ~ in elongation at break and 73 g cm/cm in tear strength. On the other hand, the membrane prepared from an aqueous mixture of collagen fiber and gelatin in the same manner as this example (Comparative Specimen D~ showed mechanical properties of: 30 in thickness, 320 kg/cm2 in tensile strength, 28 ~ in elonga-.
119~75{)7 tion at break and 65 g cm/cm in tear strength.
The facts showed that the composite material according to the present invention is excellent in strength.
EXAMPLE 8:
Aptitude test in mechanical stuffing Practical tests were carried out on the tube-form Specimen C obtained in Example 3 and the tube-form Comparative Specimen A obtained in Example 1 by making them sausage-l~ casing for a sausage meat of the following composition:
lo ll salted pork meat 1300 g ¦~ lard meat 750 g ¦l albumen 90 g ; ~¦ iced water750 g ~¦ starch 120 g - sodium glutamate 9 g allspice 12 y I¦ sucrose 9 g a phosphate 6 g pepper powder6 g -; 20 ll Table 5 shows the results when the sausage meat was ¦¦ stuffed into the two kinds of sausage-casings (Specimen C and .~ I Comparative Specimen A) using a semi-automatic stuff-worker : 760.
As is seen in Table 5, the casings prepared from a mixture of crosslinked collagen fiber and de-N-acetylated chitin tSPecimen C) were able to withstand the severe condi-. .
.
.
li~'75~)7 tions of stuffing. In addition, the knot on the sausage prepared from the sausage-casing according to the present invention (Specimen C) was smaller than that of from Compara-tive Specimen A and superior in appearance.
~I Table 5 1~
Sample Addition of Number**Stuffed Length of Rate of , de-N-acety- of run amount a sausage rupture lated chitin ~g/piece) after stuffing (cm) :.
1 yes 3 18 9.5 0 lo ~ 2 yes 9 20 9.8 0 3 yes 1 14.5 7.3 - 0
:' 11~7S~)~
., fibrous collagen and gelatin and/or soluble collagen, which may be prepared by bringing each proteinous substance into contact with the de-N-acetylated chitin, is extremely suitable to resolve the problems in the prior art.
Accordingly, the present invention provides a two-component composite comprising de-~-acetylated chitin and fibrous llcollagen wherein the amount of the de-N-acetylated chitin and ¦¦the fibrous collagen is selected from the range of 0.01 to 99 and 1 to 99.99 parts by weight, respectively, to 100 parts lo by weight of the composite material, and provides a three- or four-component composite obtained by partially replacing the : fibrous collagen of the two-component composite material with gelatin and/or soluble collagen wherein the amount of the gelatin and/or soluble collagen is up to 40 parts by weight to 100 parts by weight of the composite material and the amount . ¦ of the de-N-acetylated chitin and the fibrous collagen is the I same as in the two component composite material. In addition, the present invention provides a method for preparing the above-mentioned composite material wherein the de-N-acetylated chitin and the proteinaceous substance of the fibrous collagen, the gelatin and the soluble collagen are brought into mutual contact in an aqueous acidic medium of pH 1 to 6, preferably : pH 3 to 6 and then the obtained product is deacidified followed by cross-linking the deacidified product, if necessary.
Hitherto, proteinic composite materials such as those comprising collagen fiber and a soluble protein or comprising .' ' ' 11475~)7 collagen fiber and gelatin, etc. have been reported. However, ~I since in such a proteinic composition, the bonding between collagen and the soluble component is weak, the reduction of ~ strength of such a composition is remarkable when the composition ¦¦ is brought into hot water. On the other hand, the composite material according to the present invention does not dissolve ¦ into an aqueous salt solution, and is excellent in mechanical ¦ strength which is not reduced particularly in hot water, in non-absorbability by living bodies, in blood-coagulating lo property, in resistance against the attack of bacteria, in film-forming ability and in close adhesion to stuffed materials such as meat.
Although it has not yet been elucidated why the com-posite material according to the present invention has an excellent mechanical properties, non-abso~bability~by living bodies and a blood-coagulating property, the existence of the strong bonding via the de-N-acetylated chitin and the proteinaceous substance of the fibrous collagen, the gelatin and the soluble collagen is presumed to form a stable polyion complex as follows:
:
., ~ (3 t3 OE) (3 ~
- H3N - ~ - COO + Z-NH3 > H3N - ~ - COO H3N - Z
~:: ~9 (3 I wherein H3N-~-COO represents an ion of the proteinaceous molecule and H3N-Z represents an ion of the de-N-acety-lated chitin molecule.
., _ 5 _ `
' 11475~)7 The composite material according to the present inven-l, tion is applicable to various usages ~s one of such usages, jl an edible casing material is mentioned. Besides, as Kiriyama ~¦ et al have reported in A.N.I. No. 71, since chitosan has the ¦, following physiological activities:
Z! a) returning the raised level of serum cholesterol to normal state, b) inhibiting the raising of the level of blood sugar, c) removing growth-inhibiting substances and lo d) preventing the colonal cancer, the composite material according to the present invention may be added to foods in the form of a spherically shaped material or a fibrous material, or may exhibit the preventive or therapeutic effects to the various symptoms such as a) to d) Il as an edible material.
I Furthermore, since the composite material according ¦¦ to the present invention is not absorbable by living bodies owing to one of its components, de-N-acetylated chitin, and also has a blood-coagulating property, it can be surgically or therapeutically applied in the forms of film, fiber and those having a three dimensional structure. Although the crosslinked or not-crosslinked composite material of the present invention is itself thrombogenic, it can be applied as an anti-thrombogenic material after being heparin-treated and ~¦ possibly applied as a medical material for artificial vessels, artificial skins, artificial kidneys, etc.
-' ~ ' '' ' .
:, 11~75V7 1, , In addition, since the composite material according to the present invention is excellent in adsorbing ~roteins, ,it is possibly applicable as a base material for inmobilizing ~lenzymes and microorganisms or as an adsorbent of enzymes and ¦Imicroorganisms.
The following are the more precise description of the present invention.
The de-N-acetylated chitin mentioned in the present invention is, from the viewpoint of solubility and processability, of a degree of de-N-acetylation of 50 to 100 ~, preferably 70 to 100 % and of a viscosity of 20 to 1000 cp, the viscosity being determined at 20-C on an aqueous 1 ~ solution of acetic acid containing O.S ~ by weight of the de-N-acetylated chitin.
Co~mercialized chitosan may be used or the product obtained ¦by treating chitin in an aqueous alkali solution of~ a high concentration while heating may be used, the chitin being obtained from shells of arthropods such as crabs, lobsters, - shrimps, etc. by a conventional manner of separation and I purification.
Protein mentioned in the present invention means two ` kinds of proteins, one of which i5 the fibrous collagen in-soluble in an aqueous solvent and the other is the protein ; soluble in the aqueous solvent, i.e., gelatin and soluble collagen. They are hereinafter referred to as the proteinic component. The proteinic component is obtainable by separating ¦and purifying protelnaceous tissues from living bodies of !
11475~)7 varlous animals, or by chemically treating the separated and purified protein. The aqueous solvent mentioned in the present invention is an aqueous solution of acid or alkali. As the acid, an inorganic acid such as hydrochloric acid, etc., a single organic acid such as acetic acid, propionic acid, adipic ~ acid or a mixture of more than two kinds of the organic acid ¦Ican be exemplified. The alkali means a conventional alkali ¦¦such as sodium hydroxide, potassium hydroxide, etc.
1~ Collagen fiber has been obtained by removing non-lo collagenic components from the fibrous components consti-tuting major protein which is present in the connecting tissues of animals with chemical and mechanical treatments.
Collagen fiber, for example, may be prepared by a process wherein bovine hide or bovine Achilles tendon, after having subjected to depilating teatment, is finely cut by a mincing machine and is swollen in an aqueous acidic or alkaline medium, and then is disintegrated by a grinder to obtain ~ an aqueous dispersion, followed by filtration, if neces-I sary.
In addition, it is preferable for the improvement of strength and homogenization of the produced composite material to subject the thus finely cut hide or tendon to a pretreatment.
As a process of the pretreatment, crosslinking of amino group of lysine residue in collagen by a crosslinking agent such as formaldehyde, glutaraldehyde, dialdehyde starch, glyoxal, smoke-drying liquid, epihalohydrin, etc., succinylation . .1 . ~ ' .
.
of the amino group by succinic anhydride, acylation of the amino group by carboxylic acid anhydride or esterification of carboxyl group in the side chain of aspartic acid residue ¦lor glutamic acid residue in collagen are mentioned.
Besides, as the raw material for collagen fiber, llchips and rubbishes remaining after preparation of natural ¦¦or artificial collagen casing, which are not utllizable for ¦the casing are mentioned.
The fibrous collagen in the present invention is of 1 to 3 ~ in diameter and of 0.1 to 15 mm in length and of 5,000 to 1,000,000, preferably 20,000 to 500,000 in molecular weight, however, not necessarily being restricted wlthin such I ranges.
On the other hand, the other proteinic component of gelatin in the present invention may be one used in an indus-trial scale in food industries. The soluble collagen herein-mentioned is soluble in an aqueous solvent and may be obtained by treating the fibrous collagen with proteinase, an acid or an alkali to be wholly soluble into the aqueous solvent.
2~ Besides, a partly solubilized product obtainable in the treat-ment of the fibrous collagen is usable as a mixture of the fibrous collagen and the soluble collagen.
'I'he soluble protein of the gelatin and the soluble collagen amounts to 0 to 40 parts by weight to 100 parts by weight of the composite material according to the invention.
When the amount of the gelatin, the soluble collagen or the ., !l _ g _ .
:
I :~1475~7 mixture thereof is more than 40 parts by weight, the moisture content of the obtalned composite material is so much that the 'maintaining of the shape of the composite material is difficult and the tensile strength is poor. The gelatin is preferable as the soluble protein, and the amount of the gelatin is preferably 5 to 30 parts by weight to 100 parts by weight of the composite l! material.
¦ The soluble protein, as a component of the composite l material, contributes to the pliability, the reduced thermal lo deformation temperature, the improved mechanical strength, and also the improved dietary feeling of the composite material.
The composite material according to the present inven-l tion is also available by at first bringing the de-N-acetylated - ~ chitin and the proteinic component into contact in a medium having pH value in the range of 1 to 6, preferably,of 3 to 6 and then deacidifying of the product thus obtained followed by subjecting to crosslinking, if necessary.
The deacidifying treatment mentioned herein means the ~ adjusting of the pH of the reaction product of the de-N-acetylated ; 20 chitin and the proteinic component higher than the isoelectric point of the reaction product. The deacidificating treatment is, for instance, an addition of an alkaline solution such as sodium hydroxide, potassium hydroxide or ammonium hydroxide to bring the pH higher than 7, an addition of an aqueous solution of alkaline salt, a removal of the acid present in the reaction ~mixture by ev poration or a treatment by electrodeposition.
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li475{)7 The gelatin obtained by denaturing collagen has various isoelectric point, its jelly strength, its ash content, etc. according to the process of denaturation treatment.
~Accordingly, in the case where the proteinaceous composite ~Imaterial is prepared by an electrodeposition method from the ¦~gelatin obtained by an acid-treatment, it is necessary to adjust the pH of the electrodeposition-controlling liquid higher than 7 to make the shaped composite material on the cathode. On the other hand, in the case where the gelatin I lo obtained by alkali-treatment is used, the shaped composite material must be formed on the anode by adjusting the pH of the electrodeposition-controlling liquid lower than 7.
Whereas, in the present invention, the composite material can also be prepared by electrodeposition without the above-mentioned restriction owing to the action of the de-N-acetylated chitin which is one of the components of the composition. Meanwhile, as the gelatin for use in the case ~1~ of shaping by electrodeposition, a product containing less than 0.5 % by weight of ash, preferably less than 0.2 % by weight of ash is desirable from the viewpoints of quality and demand of electricity in preparation.
Besides, according to the process of the present I invention, by combining the proteinic component which has been insufficient in shape-forming property with the de-N-acetylated chitin, the shape-forming property, for example, membrane-forming property can be improved and the ll I
', - 11 -.
.
~1475V7 problems concerning the preparation of the composite material such as the yield in the preparation, the stability and the uniformity of the product are possibly improved. ~esides, I since the remnant free aldehyde after the pretreatment of ¦¦the fibrous collagen by the cross-linking is also reacted with ¦ the de-N-acetylated chitin according to the invention, the I composite material is desirable from the viewpoint of safety.
The following are the more detailed explanation in preparing the composite material of the present invention.
lo As one of the embodiments of the contact of the pro-teinic component and the de-N-acetylated chitin, a process comprising the steps of mixing an acid medium containing the de-N-acetylated chitin and a medium containing a proteinic component and dispersing the mixture can be exemplified. In this case, the concentration of the de-N-acetylated chitin in the acidic medium is less than 5 % by weight, preferably less than 1 % by weight in connection to its viscosity (the medium being an aqueous dilute solution of acetic acid, hydro-chloric acid, etc.), and the pH of the acidic medium is 1 to 6, preferably 3 to 6. It is not always necessary that the de-N-acetylated chitin is homogeneously dissolved in the medium, and the de-N-acetylated chitin may be dispersed uni~ormly as a fibrous state. The medium containing the proteinic com-ponent may be prepared by dispersing the proteinic component into an aqueous acidic solution of hydrochloric acid, etc.
and by adjusting the pH thereof at 3 to 6, preferably ,1 , -, 11~7507 3 to 4. However, the acidity of the medi~m is not necessarily required, that is, the medium may be neutral or alkaline. In Il this case, it is finally necessary that the p~ of the mixture ¦¦of the medium containing the de-N-acetylated chitin and the ¦medium containing the proteinic component is in the acidic ¦jregion. In addition, in the case of electrodeposition, the medium containing the proteinic component is used in an acidic ~ state.
¦ In the present invention, a film-like composite material lo is available by pouring the de-bubbled mixture of the medium containing the de-N-acetylated chitin and the medium containing the proteinic component onto a glass plate and then de-acidifying the poured mixture by the use of a drier with hot air. The film-like composite material is characterized in that its strength does not depend on the tensile direction, that is, longitudinal or transversal in spite of having the fibrous protein.
The above-mentioned characteristic feature of the film-~- like composite material according to the present invention 2~ can be exhibited by the presence of the de-N-acetylated chitin in the composite material.
I The spherically shaped composite material is prepared by dropping the de-bubbled mixture of the two media into a hydrophobic organic solvent such as toluene or xylene ` and then treating the thus ~ormed material with an aqueous alcoholic solution to deacidify the spherically shaped material.
,,1 '~' , - 13 -,.~
', 1~75~7 , .
Furthermore, fiber-like, film-like or hollowed composite material is possibly prepared by extruding the mixture fxom a hole-liXe or slit-like nozzle(s) into an aqueous solution of sodium chloride or ammonium hydroxide at a high concentration. In addition, the formed composite material of the present invention can be obtained by electrodeposition in which an aqueous medium containing the de-N-acetylated chitin and the proteinic com-ponent is introduced into an electrolytic cell provided with at least one cathode and at least one anode, and a direct voltage is loaded between both the electrodes to make the composite material accumulated on the surface of the predetermined lelectrode.
The tube-like composite material according to the present invention has an extremely thin wall rnembrane excellent in strength, and accordingly, it is highly suitable for edible casing material with a desirable dietary feeling.
In the case where the composite material according to the present invention is used for the edible casing, it is preferable that the amount of the de-N-acetylated chitin is selected from the range of 0.01 to 60 parts by weight, that of the fibrous collagen is selected from the range of 40 to 99.99 parts by weight and, if employed, that of the soluble protein of the gelatin, the soluble collagen or the mixture thereof is selected from the range of 5 to 30 parts by weight to 100 parts by weight of the composite material from the view points of the dietary feeling, the strength at smoke-drying ,,~
.
: ~ .
. ,, - 14 -.
~1~75~)7 1, and cooking, the thermal deformation temperature and the heat-stability of the casing. On the other hand, in the case where the composite material according to the present invention is ~used as a food additive, the amount of the de-N-acetylated chitin is selected from the range of 1 to 99 parts by weight to 100 parts by weight of the composite material and it is preferable to increase the amount of the de-N-acetylated chitin.
As another embodiment of the contact of the proteinic component and de-N-acetylated chitin, a process is exemplified in which a preliminarily shaped proteinic material into sheets, films, tubes, fiber-like forms, etc. is immersed into a acidified medium containing the de-N-acetylated chitin and then the shaped material is subjected to deacidifying treatment. In this case, ¦ since the superficial part of the composite materia~l is composed ¦ of the de-N-acetylated chitin c~mponent, such a shaped composite material shows extremely unique properties such as improved I mechanical properties, improved thermal resistance and anti-bacterial property.
Furthermore, in the case where the composite material according to the present invention obtained by one of the above-mentioned several methods is further subjected to a crosslinking treatment, the thus crosslinked composite material exhibits an improved property in blood-coagulation and an improved resistance to acids.
In the crosslinking treatment, a composite material : 11 . l ,' .
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il47507 according to the present invention is immersed into an a~ueous solution prepared by dissolving a crosslinking agent into a buffer solution of phosphoric acid at a pH of 7.4 for 0.1 to ll hour at 10 to 50-C. After the crosslinking is over, by ¦washing the crosslinked material with water, a water-insoluble ~crosslinked composite material is obtained.
Since it is possible by such a process of crosslinking to cause not only the crosslinking of the de-N-acetylated chitin but also the crosslinking between the de-N-acetylated chitin lo and the proteinic component, an improved bonding force in the surface of the composite material is obtained. The crosslinking occurs between the same or the different functional groups among amino groups and hydroxyl groups in the de-N-acetylated chitin and collagen. As a crosslinking agent, the same agents mentioned in the crosslinking carried out in the pretreatment ¦of the fibrous collagen are equally mentioned.
The present invention provides a novel composite material by combining the proteinic component and the de-N-`; acetylated chitin, which has excellent specific properties more than compensating the demerits of the de-N-acetylated chitin and the proteinic component, and the present invention contributes greatly to industries because the de-N-acetylated chitin is present in nature in abundance, however, owing to its high crystalinity and chemical stability, it has been utilized only in narrowly restricted fields.
,;
:' l ., , , ' - ' ' ~75()7 , ~I EXAMPLE 1 ' After immersing 100 kg of a salted steer hide into water for 15 hours, washing by water, and defatting, 90 kg of the thus treated hide was immersed into 450 kg of an aqueous liquid containing 2 ~ by weight of calcium hydroxide, 0.5 %
by weight of sodium sulfide and 0.5 % by weight of diethylamine Il at 25-C for 24 hours while gently stirring the mixture. The ¦ thus treated hide was subjected successively to a depilating I roll to remove hairs and decomposition products formed by the I treatment with calcium hydroxide, to a splitting machine to ~¦ roughly divide the hide, to a meat slicer to cut the divided I hide into tapes of 5 mm in width and then to a mincing machine to cut the tapes into 5 cm in length.
Eighty kilograms of the thus obtained hide (referred to the refined hide hereinafter) were dispersed in~o 800 kg of water, and 3 kg of acetic acid were added to the dispersion and then after stirring gently for 12 hours, the refined hide was separated from the liquid, dehydrated by centrifugation and washed with de-ionized water until the electric conductivity of the washings became less than 20~u/cm. At this time, the . ash content of the refined hide was less than 0.1 %. Forty kilograms of the thus treated refined hide were dispersed into 400 kg of an aqueous solution containing 0.015 % by weight of glutaraldehyde and adjusted to pH of 3.0 by the addition of hydrochloric acid, and then crosslinked while stirring the solution gently for 12 hours at 20C.
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11~L7507 ~~ The crosslinked refined hide was collected by filtra-tion with a wire netting and subjected to a pulp-refiner together with 113 kg of cold water to beat the fiber-bundles of the refined hide resulting in a dispersion of collagen fibers, f to which hydrochloric acid was further added to make the pH
tllereof 3Ø The thus obtained dispersion weighing 125 kg contained 2.5 % by weight of crosslinked collagen comprising a mixture of collagen fibers of 1 to 3 ~ in diameter and of 1 to 10 mm in length and still finer collagen fibriles.
lo Meanwhile, into 10 kg of an aqueous hydrochloric acid solution of pH of 3, one kg of de-N-acetylated chitin having a degree of de-N-acetylation of 95 % were dissolved and after adjusting the pH of the solution to 3.0, 20 kg of an aqueous solution of de-N-acetylated chitin were obtained.
Into 1.5 kg of the aqueous dispersion of crosslinked collagen fiber, 50 kg of de-ionized water were added and the mixture was stirred at a high speed, and then, 374 g or the : aqueous solution of de-N-acetylated chitin was added to the :~ dispersion and the mixture was stirred fGr one hour to be an ~. 20 aqueous mixture containing collagen fiber and de-N-acetylated ~:
` chitin at a weight ratio of 100/5, the viscosity and the . electric conductivity of the aqueous mixture being 200 cP
and 250 ~U /cm, respectively.
~:~ From the aqueous mixture of collagen fiber and de-N-I acetylated chitin, a tube-form membrane was formed by electro-,j - 18 -. ..: . . . . ,, , - - ,, . ~ , .
~75~)7 deposition in which a membrane was formed on the cathode (17.5 mm~) by electrophoresis of collagen fibex and de-N-acetylated chitin.
; The thus formed membrane was pulled up, at the velocity of pulling up being 10 m/min.
The thus prepared tube-form membrane just after elec-trodeposition contained 2 g of water per g of the dry membrane and showed a tensile strength of 200kg/cm2.
~¦ The pulled-up membrane, after immersing into an aqueous 7 ~ by weight of glycerol solution, was dried by a hot air at lo ¦l 75-C for 2 min, while holding the tube-form membrane by pressured air of 300 mm H2O, to be the composite material (Specimen A) - !l according to the present invention.
On the other hand, for comparison, a comparative ¦ tube-form membrane (Comparative Specimen A) consisting only of collagen fiber was prepared by the similar process mentioned above except for not adding de-N-acetylated chitin to crosslinked l collagen fiber. The Comparative Specimen A contained 7 g of ¦ water per g of the dry membrane just after electrodeposition showing a tensile strength of 100 kg/cm2.
The results of determination of tensile strength and tear strength in wet state of Specimen A and Comparative Specimen A according to the method described in JIS P-8113 and JIS P-8116 are shown in Table 1. As will be seen in Table 1, the specimen prepared from a mixture of crosslinked collagen fiber and de-N-acetylated chitin showed a remarkably improved ~ I tensile strength, and it was possible to obtain an extremely : li , ~ , .
.
~1~75V7 thin membrane.
Table 2 shows the results of strength-determination of the membranes by the same method as mentioned above after immersing into an aqueous 0.2N sodium chloride solution at 20C
for 24 hours. As will be seen in Table 2, the composite material according to the present invention is superior to the Comparative Specimen A in strength, which suggests not only the ionic bonding between collagen fiber and de-N-acetylated chitin but also another bonding factor.
.
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11475(~7 .1 .
. Table 1 ,~
Specimen Thickness Tensile Tear strength Elongation at of membrane strength (g cm/cm) ~reak (11 ) (kg/cm2) l l . ~
Specimen A 10 460 36 32 l~ Compara-ll tivè 15 250 25 25 Specimen A
. . _ _ _ ~ Table_2 . . -- - -------- -----Specimen Tensile Tear strength Elongation at strength (g cm/cm) break (kg/cm2 ) ` ( ~ ) ._ Specimen A 275 61 115 Comparative 130 48 64 ',:
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~1475V7 I EXAMPLE 2:
. . _ One kg of an acid-treated gelatin (lot number of F-795, manufactured by Miyagi Chem. Co. Ltd., Japan) was dissolved into 10 kg of an aqueous hydrochloric acid solution I ll of pH of 3, and after adjusting the pH to 3.0, 20 kg of an Il aqueous solution of gelatin was obtained.
¦ After adding 50 kg of de-ionized water to 15 kg of ¦¦ the aqueous dispersion of c?rosslinked collagen fiber prepared ¦ in Example 1 and stirring rapidly, 374 g of the aqueous lo solution of de-N-acetylated chitin prepared in Example 1 and 374 g of the aqueous solution of gelatin were further added to the dispersion of crosslinked collagen fiber, and the whole ¦ mixture was stirred for one hour to obtain an aqueous mixture of collagen fiber, de-N-acetylated chitin and gelatin at a weight ratio of collagen: de-N-acetylated chitin:, gelatin of 100 : 5 : 5.
; From the thus obtained aqueous mixture, a tube-form membrane was formed by the same method as in Example 1 (Specimen B).
For comparison, on the other hand, a comparative tube-form membrane (Comparative Specimen B) consisting of collagen and gelatin was prepared by the same process, however, without adding de-N-acetylated chitin.
The results of determination of tensile strength and tear strength of the products by the same methods described ¦in Example 1 in wet state are shown in Table 3. As is seen in ,:; ,1 , 11~7507 Il Table 3, the strength of the membrane containing collagen 1, fiber, de-N-acetylated chitin and gelatin was improved as Il compared to that of the membrane only containing collagen ¦! fiber and gelatin, thus showing the effect of added de-N-acetylated chitin.
Furthermore, the results of determination of the mechanical properties of the products after immersing them in warm water at 40-C for 24 hours shown in Table 4 indi-cate the superiority of the composite material according to the present invention in strength to the product obtained without adding de-N-acetylated chitin.
.
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. ' , Table 3 Specimen Thickness Tensile Tear strength Elongation (~) strength (g cm/cm)at break (kg/cm2) ( %) , .. _ . .. _ .
Specimen B 10 480 36 35 Specimen B 10 340 27 27 Table 4 ... . _ _ Specimen Tensile strength Tear strength .Elongation at (kg/cm2) (g cm/cm) break (~) _ Specimen B 370 60110 : Specimen B 140 4560 `.~ I
' 11~75V7 EXAMPLE 3:
A tube-form composite material (Specimen C) was pre-pared in the same manner as in Example 1 except for the weight ¦ratio of the collagen fiber to de-N-acetylated chitin of 100/20 'l and a membrane-pulling-up velocity of 25 m/min. For comparison, a sigle material (a dispersion of collagen fiber of concentra-tion of 0.58 % by weight) was processed to be a membrane, however, the membrane could not be continuously pulled up.
The mechanical properties of Specimen C were: 8 ~ in thickness, 510 kg/cm2 in tensile strength, 32 % in elongation at break and 32 g cm/cm of tear strength.
EXAMPLE 4:
A tube-form composite material (Specimen D) was prepared in the same manner as in Example 1 and 2 except for the weight ratio of collagen fiber: de-N-acetylated chitin: gelatin of 100 : 20 : 5 and a membrane-pulling-up velocity of 25 m/min.
For comparison, an aqueous mixture of dispersed collagen fiber ; and gelatin of the weight ratio of collagen fiber: gelatin of 100 : 5 was tried to be a membrane, however, the membrane could not be continuously pulled up. The mechanical properties of S~ecimen D were: 8 ~ in thickness, 490 kg/cm2, in tensile strength, 33 % in elongation at break and 33 g cm/cm in tear strength.
EXAMPLE 5:
A tube-form composite material (Specimen E) was prepared in the same manner as in Example 1 and 2 except for using . ' , ' , : ' .. ~
~ 1475~)7 llalkali-treated gelatin (lot number of F-800, Miyagi Chem. Co.
Ltd., Japan) instead of acid-treated gelatin of Example 2 and the weight ratio of collagen fiber: gelatin: de-N-acetylated l chitin of 100 : 20 : 10 and a membrane-pulling-up velocity of 15 m/min. For comparison, an aqueous mixture of collagen ~I fiber and gelatin at a weight ratio of collagen to gelatin of ¦1 100 : 20 was subjected to electrodeposition, however, no membrane deposited on the electrode. The mechanical properties of Specimen E were: 9 ~ in thickness, 440 kg/cm2 in tensile strength, 34 % in elongation at break and 30 g cm/cm in tear strength.
EXAMPLE 6:
After mixing 10 Xg of the dispersion of crosslinked collagen fiber obtained in Example 1 and 1.5 kg of an aqueous ¦ 5 % by weight of de-N-acetylated chitin solution ob,tained in Example 1 while stirring, the mixture was de-bubbled under a reduced pressure. The de-bubbled mixture was extruded from a slit of lO cm in width and of 0.3 mm in height into an aqueous O.lN ammonium hydroxide solution contained in a coagulation bath. The thus obtained film-form material was washed with water, and was treated in the same manner as in Example 1 to be a film-form composite material according to the present invention (Specimen F).
The results of the determination in a manner as in Example 1 were: 30 ~ in thickness, 520 kg/cm2 in tensile stren~th, 30 % in elongation at break and 75 g cm/cm of tear strength.
`::
~1~75{)7 On the other hand, a film prepared from the same , dispersion of crosslinked collagen fiber in the same manner as above (Comparative Specimen C) without adding the solution of de-N-acetylated chitin showed the following properties:
30 ~ in thickness, 200 kg/cm2 in tensile strength, 26 % in elongation at break and 48 g cm/cm in tear strength.
From the above-mentioned results, the composite material ¦ according to the present invention was confirmed to be superior to the single material (only consisting of crosslinked collagen fiber) in mechanical strength.
EXAMPLE 7:
; After mixing 10 kg of the dispersion of crosslinked collagen fiber obtained in Example 1, 1 kg of an aqueous 5 %
by weight gelatin solution obtained in Example 2 and 1.5 kg ;~ of an aqueous 5 % by weight de-N-acetylated chitin solution obtained in Example 1, the mixture was well stirred and de-bubbled under a reduced pressure, and subjected to the same ¦ procedures as in Example 6 to obtain a film-form composite material according to the present invention (Specimen G).
The mechanical properties of Specimen G in wet state were:
30 ~ in thickness, 495 kg/cm in tensile strength, 31 ~ in elongation at break and 73 g cm/cm in tear strength. On the other hand, the membrane prepared from an aqueous mixture of collagen fiber and gelatin in the same manner as this example (Comparative Specimen D~ showed mechanical properties of: 30 in thickness, 320 kg/cm2 in tensile strength, 28 ~ in elonga-.
119~75{)7 tion at break and 65 g cm/cm in tear strength.
The facts showed that the composite material according to the present invention is excellent in strength.
EXAMPLE 8:
Aptitude test in mechanical stuffing Practical tests were carried out on the tube-form Specimen C obtained in Example 3 and the tube-form Comparative Specimen A obtained in Example 1 by making them sausage-l~ casing for a sausage meat of the following composition:
lo ll salted pork meat 1300 g ¦~ lard meat 750 g ¦l albumen 90 g ; ~¦ iced water750 g ~¦ starch 120 g - sodium glutamate 9 g allspice 12 y I¦ sucrose 9 g a phosphate 6 g pepper powder6 g -; 20 ll Table 5 shows the results when the sausage meat was ¦¦ stuffed into the two kinds of sausage-casings (Specimen C and .~ I Comparative Specimen A) using a semi-automatic stuff-worker : 760.
As is seen in Table 5, the casings prepared from a mixture of crosslinked collagen fiber and de-N-acetylated chitin tSPecimen C) were able to withstand the severe condi-. .
.
.
li~'75~)7 tions of stuffing. In addition, the knot on the sausage prepared from the sausage-casing according to the present invention (Specimen C) was smaller than that of from Compara-tive Specimen A and superior in appearance.
~I Table 5 1~
Sample Addition of Number**Stuffed Length of Rate of , de-N-acety- of run amount a sausage rupture lated chitin ~g/piece) after stuffing (cm) :.
1 yes 3 18 9.5 0 lo ~ 2 yes 9 20 9.8 0 3 yes 1 14.5 7.3 - 0
4 yes 1 17.5 8.4 0 yes 1 20 9.9 0 6 yes 1 23.511.8 0 __ 7 no 1 17.2 9.5 o 8 no 2 19.410.5 1/400 9 no 1 15.4 7.4 1/250 Notes: (*) Sample No. 1 to 6 used Specimen C according to the present invention. Sample No. 7 to 9 used Com-parative Specimen A.
(**)By one run the tube-form material of 1000 m is produced in the electrodeposition procedure.
; - 29 -- ` - , .
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11~75{17 . E~IPLE 9:
_ Aptitude test in mechanical sturfing ' The same practical tests were carried out on the tube-,. formed Specimen D obtained in Example 4 and Comparative Specimen B obtained in Example 2 as sausage-casings with the same ' sausage meat as in Example 8. Table 6 shows the results of ,' stuffing when the sausage meat was stuffed into the two kinds ! of sausage-casings (Specimen D and Comparative Specimen B) while using a semi-automatic stuff-worker 760.
lO llAs is seen in Table 6, the casings prepared from an aqueous mixture of dispersed collagen fiber and de-N-acetylated chitin and gelatin (Specimen D) according to the present invention were able to withstand the severe conditions of stuffing. In addition, the knot on the sausages prepared 1¦ from the sausage-casing according to the present i~vention ., I
~Specimen D) was smaller than that of Comparative Specimen ¦ B and superior also in appearance.
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Table 6 ll Sample Addition of Number of stuîfed Length of Rate il No. de-N-acety- run amountsausage of , lated chitin (g/piece) (cm) rupture l l 1, 1 yes 3 18 9.5 0 , 2 yes 9 20 9.8 0 : I 3 yes 1 14.57.3 0 4 yes 1 17.58.4 0 yes 1 20 9.9 0 .
6 yes 1 23.511.8 0 . ~
7 no 1 17.29.5 0 8 no 2 19.410.5 1/1000 9 no~ 1 15.47.4 1/500 EXAMPLE 10:
_ _ _ Sausages of Sample No. 2 and No. 8 prepared in Example 8 were smoked as follows:
Pieces of sausage were hanged from a bar with eight rings in a chain-like state, and put into a smoking chamber.
They were dried for 30 min at a controlled temperature of 55 to 60-C at first, and then the temperature was raised to 75-C
¦ while introducing a smoke generated from wood chips and further I I
' .
- - , il~75{)7 introduciny steam into the chamber to control the humidity of the chamber at a constant level, the smoking treatment being continued for 30 min.
Then, steam was again introduced into the chamber to raise the temperature to 75 to 80-C and after keeping the temperature for 50 min, the sausages were taken out from the chamber and cold water was sprayed onto the sausages to quench them.
The thus treated sausages were subjected to the follow-~ lo ¦ ing rupture tests under the simulated conditions of cooking of the sausages while supposing the cooking in homes by house-wlves:
l) Fry-test Into a frying pan provided with a temperature controller, an oil for frying was placed and while keeping the1temperature of the oil at 160-C, every two sausages were put into the pan at a time in total 5 times (for preventing the reduction of the oil temperature), and the number of rupture of the ten sausages within 30 sec/time were counted.
2) Frying pan test A frying pan was set so that the bottom of the pan was immersed in an oil bath and was lower than the surface of the oil by 10 mm, and the temperature of the oil ba~th was controlled so that the surface of the pan was kept at 175 to 180~C.
Then, a small amount of the oil for frying was painted on the whole surface of the pan and every five sausages of the l!
.
~1475~)7 ' prepared ten sausages were put into the pan and moved within the pan slowly for 2 min, and the number of rupture of the sausage was counted. The test was carried out two times on the same specimen.
3) Boiling test While keeping water in a pot of 50 mm in depth at boiling, every five sausages of the prepared ten sausages were Il put into the pot and the number of rupture of the sausage within ¦¦ 5 min of boiling was counted. The test was carried out two lo times on the same Sample.
The results of the three kinds of tests are shown in Table 7.
Table 7 .. . . ................... ._. ._ .
I Sample Addition of Number of rupture per lQ sausages No. de-N-acety-lated chitin Fry-test Frying pan test Boiling test ..._ ._ ... ..
2 yes 1 0 1 .. _ .
8 no 5 2 10 As is seen in Table 7, the sausage-casing of the material according to the present invention showed an excellent results also in practical cooking tests.
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, 75~)7 In order to support these practical facts, the respec-tive membranes of these two kinds of sausage casing were immersed into a hot water at 75C for one min, and the percentage of their shrinkage and the thermal deformation temperature were determined. The results are shown in Table 8.
Table 8 Il .__ ._ __ .. , Specimen Addition of Thermal Percent shrinkage I Il de-N-acety- deformation I lated chitin temperature lo (C) longltudLnal transversal direction direction . . . .. _ _ _ Specimen yes 54.5 8.5 10.5 ...
Compara-~o 52 35.3 3.3 .
I Besides, the sausage with the casing prepared from Icrosslinked collagen fiber added with de-N-acetylated chitin gives a feeling of mellow taste and sweetness when taken :~: into one's mouth not hitherto have been felt on other sausages, for instance, with a membrane prepared from collagen fibers only.
I I
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In the same manner as in Example 1, tube-form material~ having weight ratios of collagen fiber to de-N-acetylated chitin of 100/1 and 100/0.1, respectively, were prepared.
,Into these tube-form materials, meat mixture was ;I stuffed under the conditions of Sample No. 2 of Example 8 ¦ to be sausages, and the sausages were smoked in the same manner il as in Example 10.
lo ¦¦ No rupture of the sausages was observed in stuffing, I and the results of the cooking ~ests carried out as in Example ¦ 10 are shown in Table 9.
~ Table 9 I
Amount of de-N-acetylated Number of rupture per 10 sausage chitin added per 100 parts by weight of collagen fiber Test l* Test 2** Test 3 ***
. ~.. _ 1 0 . _.
0.1 2 0 2 Notes: Test l* : Fry-test, Test 2**: Frying pan test, and Test 3***: Boiling test As is seen in Table 9, the composite material accord-ing to the present invention showed an excellent result even in practical cooking tests.
1!
; - 35 -, 119~7507 Xi~1PLE 12:
Sausayes of Sample No. 2 and No. 8 prepared in . Example 9 were smoked in the same manner as in ~xample 10.
The smoked sausages were subjected to the rupture tests under . the simulated conditions of cooking of the sausages while Il supposing the cooking in homes by housewives as in Example j! lo. The results of the three kinds of tests are shown in ~¦ Table 10.
Table 10 . .~
lo Sample Addition ofNumber of rupture per 10 sausages No.de-N-acetylated _ _ chltin _ Fry-test Frying pan-test Boiling-2 yes 1 0 3 . ._ .
8 no ¦ 3 2 ~ 10 As is seen in Table 10, the sausage-casing of the composite material according to the present invention showed an excellent results also in practical cooking tests.
Besides, when the thus prepared sausage from the aqueous mixture containing also de-N-acetylated chitin was ~ 20 taken into the consumer's mouth, a mellow taste and sweetness : spreaded throughout the mouth giving an unparalleled feeling of eating never given by the sausages using only collagen as the casing material. In the sensory tests by a panel consisting I
11475()7 of 30 persons, a general evaluation that no artificial feeling was qiven by the sausage stuffed in the casing of the composite material o~ the present invention as compared to the sausage stuffed into natural sheep gut was obtained.
EXAMPLE 13:
Into 100 litres of acetic anhydride at a temperature of 15'C, 50 kg of the refined hide dehydrated by centrifuging in Example 1 were immersed for 8 hours to have collagen fiber acetylated. The thus treated refined hide was washed with flowing de-ionized water. The isoelectric point of the original steer hide of pH of 6.5 changed to 3.8 by the acetylation.
An aqueous dispersion of collagen fibers was prepared from the acetylated hide in the same manner as in Example 1, the content f collagen being 2.5 ~. -An aqueous solution of de-N-acetylated chitin obtained ¦in Example 1 was added to the thus prepared aqueous dispersion of collagen fiber in the same manner as in Example 1, however, with the ratio of collagen to de-N-acetylated chitin of 100/10 to prepare an aqueous mixture of collagen fiber and de-N-acetylated chitin.
An electrodeposited membrane was prepared from the aqueous mixture as in Example 1 with the same a~ter-treatment as in Example 1.
The mechanical properties of the thus prepared tube-form membrane determined as in Example 1 were: 9 ~ in thickness, 485 kg/cm2 in tensile strength, 33 ~ in elongation at brea~
, ~475~)7 ,i i.
and 35 g cm/cm in tear strength.
EXAMPLE 14:
Using the refined hide dehydrated by centrifuging in the same manner as in Example 1, a 2.5 ~ dispersion of collagen in water of pH of 3.0 was prepared as in Example 13, and then the aqueous solution of de-N-acetylated chitin obtained ~¦ in Example 1 and the aqueous solution of gelatin obtained in ¦ Example 2 were added to the dispersion to obtain an aqueous mixture of dispersed collagen fiber and gelatin and de-N-lo acetylated chitin at a weight ratio of 100:10:10. The thus prepared aqueous mixture was subjected to electrodeposition in the same manner as in Example 1 to prepare a tube form membrane, one of the composite materials of the present invention.
The mechanical properties of the membrane determined as in Example 1 were: 9 ~ in thickness, 480 kg/cm2~in tensile strength, 33 ~ in elongation at break and 34 g cm/cm in tear strength.
EXAMPLE 15:
~ Into 20 litres of Atkins-Pantin buffer solution of pH
1~ 20 of 9 containing 0.2 ~ by weight of "Pronasel' (manufactured by Kaken Chem. Co. Ltd., Japan), one kg of the refined hide obtained in Example 1 before crosslinking treatment was immersed for 24 hours under slow stirring to bring the hide into reac-tion with Pronase. After the reaction was over, the lumpy material was recovered by centrifugation and washed with water to purify. The purified material was dissolved into an aqueous lN hydrochloric acid solution of pH of 3.5 to obtain 100 kg of an uniform solution of solubilized collagen. On the determination * Trade Mark - ~ ' ~ 75~7 of the concentration of solubilized collagen in the solution by taking and examining a small portion of the solution after ll freeze-drying, it was known to be 0.95 % by weight.
¦I To 13.5 kg of the aqueous 2.5 % crosslinked collagen ¦~ dispersion prepared in Example 1, 50 kg of de-ionized water were added under high speed stirring, and 3.95 kg of the thus obtained aqueous solution of solubilized collagen were added to the mixture, and then the whole mixture was stirred for 30 min.
lo To the thus prepared mixture, 748 g of the aqueous de-N-acetylated chitin solution prepared in ~xample 1 and 374 g of the aqueous gelatin solution prepared in Example 2 were added, and the mixture was stirred well. In the thus prepared aqueous mixture, the weight ratio of collagen fibers: soluble collagen: gelatin: de-N-acetylated chitin was 90~: 10 : 10 : 20.
By subjecting the aqueous mixture to electrodeposition while using the apparatus used in Example 1, a composite material, i.e., a tube-form membrane, of the present invention was obtained of which the mechanical properties, determined as in Example 1, were: 9 ~-in thickness, 500 kg/cm2 in tensile strength, 35 % in elongation at break and 35 g cm/cm in tear strength.
EXAMPLE 16:
Into 50 g of an aqueous dispersion of 2.5 % by weight ` of not-crosslinked collagen prepared following the procedures in Example 1, one gram of acidic protease was added, and after keeping the mixture for 5 hours at 37C, , I I
1~475Q7 l; :
the mixture was neutralized, subjected to centrifugation and the solid matter recovered was washed with water, and then adjusted to pH of 3.5 by the addition o' an aqueous lN hydro-chloric acid solution. The thus prepared mixture weighing 1 kg was subjected to homogenization while Xeeping at lower than lO~C to be further finely divided. The thus obtained liquid Il is hereinafter referred to as Liquid B.
¦¦ A homogeneous solution of 20 g of de-N-acetylated ~ chitin dissolved in an aqueous 5 % by weight of acetic acid lo I solution, hereinafter referred to as ~iquid A, was added to ¦ Liquid B, and the mixture was processed to be a homogeneous solution by a homogenizer.
Into a 3-litre flask provided a stirrer, in which 2 litres of decahydronaphthalene and one gram of polyoxyethylene ¦ sorbitan (trade mark ~Twin 80) were placed, 100 ml of the ¦ mixture of Liquid A and Liquid B was added, and the whole mixture was stirred at 1,000 rpm for one hour to be a disper-sion. By re-dispersing the dispersion in 10 litres of ethanol, an insoluble matter was obtained. The insoluble matter was ; 20 recovered by filtration and washed repeatedly with ethanol to remove the organic substances except ethanol, and the thus treated insoluble substance was re-dispersed into one litre of an aqueous 2 % ammonium hydroxide solution. After neutrali-zing the dispersion, and recovering the precipitate by filtra-¦ tion and washing with water, spherical particles of a composite material according to the present invention were obtained ;~' ' '', ~1475~)7 (Specimen H) of particle size of 0.1 to l.0 mm in diameter.
After freeze-drying Specimen H, one gram of dried Specimen H was put into an aqueous physiological saline solu-tion, the solution was filtered off to recover the spherical particles.
The thus treated spherical particles were filled into a glass column of 8 mm in internal diameter, and then 50 ml of rabbit's blood added with 250 units of heparin were perfused t~rough the column at a rate of 20 ml/min at 37-C
for 15 min. After stopping the circulation, the blood was removed from the column as soon as possible, and the column was washed with an aqueous physiological saline solution, and then the spherical particles were taken out from the column to examine the degree of adherence of platelets and blood corpuscles on the particles. The adherence of both the platelets and blood corpuscles was clearly recognized.
EXAMPLE 17:
To 50 g of the aqueous dispersion of crosslinked collagen fiber prepared in Example l, 7.5 g of the aqueous solution of gelatin of Example 2 were added, and after adjusting the pH of the mixture by the addition of an lN aqueous hydro-chloric acid solution to 3.5, the mixture amounting to l kg was subjected at a temperature of lower than lO'C to a homo-genizer to be finely divided collagen therein, the thus prepared dispersion being referred to as Liquid C.
A mixture of Liquid C and Liquid A of Example 16 was ,.
; - 41 -~ , , . ' .
~1475V7 prepared and subjected to homogenization to obtain a homo-, geneous dispersion.
Into a 3-litre flask provided witn a stirrer, in which ~1 2 litres of decahydronaphthalene and one gram of polyox~ethylene I sorbitan (Trade mark ~Twin #80) were placed, 100 ml of the ,,lhomogeneous dispersion of the mixture of Liquids C and A were introduced, and after one hour of stirring at 1000 rpm, the ~mixture was re-dispersed into 10 litres of ethanol to obtain an insoluble material. The insoluble material was collected by filtration and washed repeatedly with ethanol to remove decahydronaphthalene, and then, the washed material was again dispersed into one litre of an aqueous 2 % ammonium hydroxide solution. On neutralizing the dispersion, spherical particles were deposited. The particles were collected by filtration, and washed with water to obtain a composite material o~ the present invention (Specimen K) as spherical particles of 0.1 to 1.0 mm in diameter.
Results of examination of the surface of the particles of Specimen K as in Example 16 showed the adherence of platelets and blood corpuscles on the surface of the particles, Specimen X.
EXAMPLE 18:
After pouring the aqueous mixture of collagen fiber and de-N-acetylated chitin prepared in Example 6 onto a plate :¦~ of 200 mm square to a thickness of 5 mm and leaviny still for .
I one hour, the plate with the mixture was immersed into an ¦aqueous 0.lN ammonium hydroxide solution to neutralize the 1,1 ,1 ; .
'' " ` ''.
1 11475~7 1, .
Il acidity, and washed with de-ionized water to remove salts.
,~~ The thus obtained material was immersed into an aqueous 0.1 M
disodium phosphate solution containing 0.5 % by weight of form- ¦
aldehyde at 30C for one hour and then washed with water.
The thus obtained formed material was freeze-dried to be a sponge-form material. The sponge-form material Il showed an absorption of aqueous physiological saline solution ¦ of 20 g per g of the material and further, it shows a strong coagulation activity to bloods, and accordingly, it has been lo found that the material is possib~'ly applicable as a menstrual pad material and as a garrot tourniquet for urgent cases.
EXAMPLE l9:
~ The aqueous mixture of collagen fiber, gelatin and ¦I de-N-acetylated chitin of Example 2 was treated in the same manner as in Example 18 to obtain a spongy-form ma~erial. The thus obtained material showed an absorption ability to an aqueous physiological saline solution of 21 g/g material, and a strong ~`~ coagulating property to human blood. Accordingly, it was found that the material is applicable as the material for menstrual pads and tourniquets for urgent cases.
EXAMPLE 20:
Into a solution prepared by dissolving 1 g of glucose-~ isomerase(2000 U/g; manufactured by Nagase Sangyo Co. Ltd.) I into an aqueous O.lM disodium phosphate solution, 30 g of the spherical particles (Specimen ~ of Example 16) were added, and ; I after stirring the mixture for 2 hours at 5~C, 2 ml of an , .
' " ' ', ` ~1475~7 aqueous 25 % by weight glutaraldehyde solution was added to the mixture. After leaving the mixture as it is for 5 hours, solid materials were recovered by filtration, washed with an aqueous phosphoric buffer solution of pH of 7.0 and added to S00 ml of an aqueous phosphoric buffer solution containing 2 g of glucose. After thus treating the mixture for 60 min at 70-C, the contents of glucose and fructose of the solid material were 600 and 1400 mg, respectively, together with the fixed enzyme I (glucose-isomerase) of 70 ~ of the applied amount.
lo In addition, separately, the pH of the solution of collagen and de-~-acetylated chitin before re-dispersion in il Example 16 was adjusted to 6.0, and 1 g of glucose-isomerase ~ was added to the solution at 5-C. After re-dispersing the ¦I mixture as in Example 16, the solid matter was recovered by ¦ filtration and washed with ethanol and then with an aqueous phosphoric buffer solution of pH of 7Ø
It was found that 85 ~ of the enzyme (glucose-isomerase) applied was fixed onto the solid matter.
These results show that the thus obtained material is possibly applied as a carrier for fixed enzymes and microbial bodies.
EXAMPLE 21:
The same procedures as in Example 20 were carried out ; except for using Specimen X obtained in Example 17 instead of Specimen H used in Example 20. The amounts of glucose and ¦ fructose on Specimen K after the treatment at 70-C for 60 min '.~, , - 44 -, 1 ~1475()7 ., were 600 and 1400 mg, respectively with the fixed rate of glucose isomerase of 70 ~.
¦ From these results, it has been found that a compo-site material according to the present invention, i.e., Specimen K, is possibly applicable as a carrier of fixed enzymes and microbial bodies.
EXAMPLE 22: ~
Ater immersing Comparative Specimen A comprising a shaped material consisting only of collagen fibers into a lo homogeneous solution of a viscosity of 50 cP prepared by adding
(**)By one run the tube-form material of 1000 m is produced in the electrodeposition procedure.
; - 29 -- ` - , .
- '-.
11~75{17 . E~IPLE 9:
_ Aptitude test in mechanical sturfing ' The same practical tests were carried out on the tube-,. formed Specimen D obtained in Example 4 and Comparative Specimen B obtained in Example 2 as sausage-casings with the same ' sausage meat as in Example 8. Table 6 shows the results of ,' stuffing when the sausage meat was stuffed into the two kinds ! of sausage-casings (Specimen D and Comparative Specimen B) while using a semi-automatic stuff-worker 760.
lO llAs is seen in Table 6, the casings prepared from an aqueous mixture of dispersed collagen fiber and de-N-acetylated chitin and gelatin (Specimen D) according to the present invention were able to withstand the severe conditions of stuffing. In addition, the knot on the sausages prepared 1¦ from the sausage-casing according to the present i~vention ., I
~Specimen D) was smaller than that of Comparative Specimen ¦ B and superior also in appearance.
.,, I
: - 30 -, `~
'.
Table 6 ll Sample Addition of Number of stuîfed Length of Rate il No. de-N-acety- run amountsausage of , lated chitin (g/piece) (cm) rupture l l 1, 1 yes 3 18 9.5 0 , 2 yes 9 20 9.8 0 : I 3 yes 1 14.57.3 0 4 yes 1 17.58.4 0 yes 1 20 9.9 0 .
6 yes 1 23.511.8 0 . ~
7 no 1 17.29.5 0 8 no 2 19.410.5 1/1000 9 no~ 1 15.47.4 1/500 EXAMPLE 10:
_ _ _ Sausages of Sample No. 2 and No. 8 prepared in Example 8 were smoked as follows:
Pieces of sausage were hanged from a bar with eight rings in a chain-like state, and put into a smoking chamber.
They were dried for 30 min at a controlled temperature of 55 to 60-C at first, and then the temperature was raised to 75-C
¦ while introducing a smoke generated from wood chips and further I I
' .
- - , il~75{)7 introduciny steam into the chamber to control the humidity of the chamber at a constant level, the smoking treatment being continued for 30 min.
Then, steam was again introduced into the chamber to raise the temperature to 75 to 80-C and after keeping the temperature for 50 min, the sausages were taken out from the chamber and cold water was sprayed onto the sausages to quench them.
The thus treated sausages were subjected to the follow-~ lo ¦ ing rupture tests under the simulated conditions of cooking of the sausages while supposing the cooking in homes by house-wlves:
l) Fry-test Into a frying pan provided with a temperature controller, an oil for frying was placed and while keeping the1temperature of the oil at 160-C, every two sausages were put into the pan at a time in total 5 times (for preventing the reduction of the oil temperature), and the number of rupture of the ten sausages within 30 sec/time were counted.
2) Frying pan test A frying pan was set so that the bottom of the pan was immersed in an oil bath and was lower than the surface of the oil by 10 mm, and the temperature of the oil ba~th was controlled so that the surface of the pan was kept at 175 to 180~C.
Then, a small amount of the oil for frying was painted on the whole surface of the pan and every five sausages of the l!
.
~1475~)7 ' prepared ten sausages were put into the pan and moved within the pan slowly for 2 min, and the number of rupture of the sausage was counted. The test was carried out two times on the same specimen.
3) Boiling test While keeping water in a pot of 50 mm in depth at boiling, every five sausages of the prepared ten sausages were Il put into the pot and the number of rupture of the sausage within ¦¦ 5 min of boiling was counted. The test was carried out two lo times on the same Sample.
The results of the three kinds of tests are shown in Table 7.
Table 7 .. . . ................... ._. ._ .
I Sample Addition of Number of rupture per lQ sausages No. de-N-acety-lated chitin Fry-test Frying pan test Boiling test ..._ ._ ... ..
2 yes 1 0 1 .. _ .
8 no 5 2 10 As is seen in Table 7, the sausage-casing of the material according to the present invention showed an excellent results also in practical cooking tests.
,'. .
, ' ~. , .
' ' ' ', ' ':' ' ' ' ' ' ' ' ' . . . .
, 75~)7 In order to support these practical facts, the respec-tive membranes of these two kinds of sausage casing were immersed into a hot water at 75C for one min, and the percentage of their shrinkage and the thermal deformation temperature were determined. The results are shown in Table 8.
Table 8 Il .__ ._ __ .. , Specimen Addition of Thermal Percent shrinkage I Il de-N-acety- deformation I lated chitin temperature lo (C) longltudLnal transversal direction direction . . . .. _ _ _ Specimen yes 54.5 8.5 10.5 ...
Compara-~o 52 35.3 3.3 .
I Besides, the sausage with the casing prepared from Icrosslinked collagen fiber added with de-N-acetylated chitin gives a feeling of mellow taste and sweetness when taken :~: into one's mouth not hitherto have been felt on other sausages, for instance, with a membrane prepared from collagen fibers only.
I I
' i.
, -:
, ''~, EXAMPLE 11:
In the same manner as in Example 1, tube-form material~ having weight ratios of collagen fiber to de-N-acetylated chitin of 100/1 and 100/0.1, respectively, were prepared.
,Into these tube-form materials, meat mixture was ;I stuffed under the conditions of Sample No. 2 of Example 8 ¦ to be sausages, and the sausages were smoked in the same manner il as in Example 10.
lo ¦¦ No rupture of the sausages was observed in stuffing, I and the results of the cooking ~ests carried out as in Example ¦ 10 are shown in Table 9.
~ Table 9 I
Amount of de-N-acetylated Number of rupture per 10 sausage chitin added per 100 parts by weight of collagen fiber Test l* Test 2** Test 3 ***
. ~.. _ 1 0 . _.
0.1 2 0 2 Notes: Test l* : Fry-test, Test 2**: Frying pan test, and Test 3***: Boiling test As is seen in Table 9, the composite material accord-ing to the present invention showed an excellent result even in practical cooking tests.
1!
; - 35 -, 119~7507 Xi~1PLE 12:
Sausayes of Sample No. 2 and No. 8 prepared in . Example 9 were smoked in the same manner as in ~xample 10.
The smoked sausages were subjected to the rupture tests under . the simulated conditions of cooking of the sausages while Il supposing the cooking in homes by housewives as in Example j! lo. The results of the three kinds of tests are shown in ~¦ Table 10.
Table 10 . .~
lo Sample Addition ofNumber of rupture per 10 sausages No.de-N-acetylated _ _ chltin _ Fry-test Frying pan-test Boiling-2 yes 1 0 3 . ._ .
8 no ¦ 3 2 ~ 10 As is seen in Table 10, the sausage-casing of the composite material according to the present invention showed an excellent results also in practical cooking tests.
Besides, when the thus prepared sausage from the aqueous mixture containing also de-N-acetylated chitin was ~ 20 taken into the consumer's mouth, a mellow taste and sweetness : spreaded throughout the mouth giving an unparalleled feeling of eating never given by the sausages using only collagen as the casing material. In the sensory tests by a panel consisting I
11475()7 of 30 persons, a general evaluation that no artificial feeling was qiven by the sausage stuffed in the casing of the composite material o~ the present invention as compared to the sausage stuffed into natural sheep gut was obtained.
EXAMPLE 13:
Into 100 litres of acetic anhydride at a temperature of 15'C, 50 kg of the refined hide dehydrated by centrifuging in Example 1 were immersed for 8 hours to have collagen fiber acetylated. The thus treated refined hide was washed with flowing de-ionized water. The isoelectric point of the original steer hide of pH of 6.5 changed to 3.8 by the acetylation.
An aqueous dispersion of collagen fibers was prepared from the acetylated hide in the same manner as in Example 1, the content f collagen being 2.5 ~. -An aqueous solution of de-N-acetylated chitin obtained ¦in Example 1 was added to the thus prepared aqueous dispersion of collagen fiber in the same manner as in Example 1, however, with the ratio of collagen to de-N-acetylated chitin of 100/10 to prepare an aqueous mixture of collagen fiber and de-N-acetylated chitin.
An electrodeposited membrane was prepared from the aqueous mixture as in Example 1 with the same a~ter-treatment as in Example 1.
The mechanical properties of the thus prepared tube-form membrane determined as in Example 1 were: 9 ~ in thickness, 485 kg/cm2 in tensile strength, 33 ~ in elongation at brea~
, ~475~)7 ,i i.
and 35 g cm/cm in tear strength.
EXAMPLE 14:
Using the refined hide dehydrated by centrifuging in the same manner as in Example 1, a 2.5 ~ dispersion of collagen in water of pH of 3.0 was prepared as in Example 13, and then the aqueous solution of de-N-acetylated chitin obtained ~¦ in Example 1 and the aqueous solution of gelatin obtained in ¦ Example 2 were added to the dispersion to obtain an aqueous mixture of dispersed collagen fiber and gelatin and de-N-lo acetylated chitin at a weight ratio of 100:10:10. The thus prepared aqueous mixture was subjected to electrodeposition in the same manner as in Example 1 to prepare a tube form membrane, one of the composite materials of the present invention.
The mechanical properties of the membrane determined as in Example 1 were: 9 ~ in thickness, 480 kg/cm2~in tensile strength, 33 ~ in elongation at break and 34 g cm/cm in tear strength.
EXAMPLE 15:
~ Into 20 litres of Atkins-Pantin buffer solution of pH
1~ 20 of 9 containing 0.2 ~ by weight of "Pronasel' (manufactured by Kaken Chem. Co. Ltd., Japan), one kg of the refined hide obtained in Example 1 before crosslinking treatment was immersed for 24 hours under slow stirring to bring the hide into reac-tion with Pronase. After the reaction was over, the lumpy material was recovered by centrifugation and washed with water to purify. The purified material was dissolved into an aqueous lN hydrochloric acid solution of pH of 3.5 to obtain 100 kg of an uniform solution of solubilized collagen. On the determination * Trade Mark - ~ ' ~ 75~7 of the concentration of solubilized collagen in the solution by taking and examining a small portion of the solution after ll freeze-drying, it was known to be 0.95 % by weight.
¦I To 13.5 kg of the aqueous 2.5 % crosslinked collagen ¦~ dispersion prepared in Example 1, 50 kg of de-ionized water were added under high speed stirring, and 3.95 kg of the thus obtained aqueous solution of solubilized collagen were added to the mixture, and then the whole mixture was stirred for 30 min.
lo To the thus prepared mixture, 748 g of the aqueous de-N-acetylated chitin solution prepared in ~xample 1 and 374 g of the aqueous gelatin solution prepared in Example 2 were added, and the mixture was stirred well. In the thus prepared aqueous mixture, the weight ratio of collagen fibers: soluble collagen: gelatin: de-N-acetylated chitin was 90~: 10 : 10 : 20.
By subjecting the aqueous mixture to electrodeposition while using the apparatus used in Example 1, a composite material, i.e., a tube-form membrane, of the present invention was obtained of which the mechanical properties, determined as in Example 1, were: 9 ~-in thickness, 500 kg/cm2 in tensile strength, 35 % in elongation at break and 35 g cm/cm in tear strength.
EXAMPLE 16:
Into 50 g of an aqueous dispersion of 2.5 % by weight ` of not-crosslinked collagen prepared following the procedures in Example 1, one gram of acidic protease was added, and after keeping the mixture for 5 hours at 37C, , I I
1~475Q7 l; :
the mixture was neutralized, subjected to centrifugation and the solid matter recovered was washed with water, and then adjusted to pH of 3.5 by the addition o' an aqueous lN hydro-chloric acid solution. The thus prepared mixture weighing 1 kg was subjected to homogenization while Xeeping at lower than lO~C to be further finely divided. The thus obtained liquid Il is hereinafter referred to as Liquid B.
¦¦ A homogeneous solution of 20 g of de-N-acetylated ~ chitin dissolved in an aqueous 5 % by weight of acetic acid lo I solution, hereinafter referred to as ~iquid A, was added to ¦ Liquid B, and the mixture was processed to be a homogeneous solution by a homogenizer.
Into a 3-litre flask provided a stirrer, in which 2 litres of decahydronaphthalene and one gram of polyoxyethylene ¦ sorbitan (trade mark ~Twin 80) were placed, 100 ml of the ¦ mixture of Liquid A and Liquid B was added, and the whole mixture was stirred at 1,000 rpm for one hour to be a disper-sion. By re-dispersing the dispersion in 10 litres of ethanol, an insoluble matter was obtained. The insoluble matter was ; 20 recovered by filtration and washed repeatedly with ethanol to remove the organic substances except ethanol, and the thus treated insoluble substance was re-dispersed into one litre of an aqueous 2 % ammonium hydroxide solution. After neutrali-zing the dispersion, and recovering the precipitate by filtra-¦ tion and washing with water, spherical particles of a composite material according to the present invention were obtained ;~' ' '', ~1475~)7 (Specimen H) of particle size of 0.1 to l.0 mm in diameter.
After freeze-drying Specimen H, one gram of dried Specimen H was put into an aqueous physiological saline solu-tion, the solution was filtered off to recover the spherical particles.
The thus treated spherical particles were filled into a glass column of 8 mm in internal diameter, and then 50 ml of rabbit's blood added with 250 units of heparin were perfused t~rough the column at a rate of 20 ml/min at 37-C
for 15 min. After stopping the circulation, the blood was removed from the column as soon as possible, and the column was washed with an aqueous physiological saline solution, and then the spherical particles were taken out from the column to examine the degree of adherence of platelets and blood corpuscles on the particles. The adherence of both the platelets and blood corpuscles was clearly recognized.
EXAMPLE 17:
To 50 g of the aqueous dispersion of crosslinked collagen fiber prepared in Example l, 7.5 g of the aqueous solution of gelatin of Example 2 were added, and after adjusting the pH of the mixture by the addition of an lN aqueous hydro-chloric acid solution to 3.5, the mixture amounting to l kg was subjected at a temperature of lower than lO'C to a homo-genizer to be finely divided collagen therein, the thus prepared dispersion being referred to as Liquid C.
A mixture of Liquid C and Liquid A of Example 16 was ,.
; - 41 -~ , , . ' .
~1475V7 prepared and subjected to homogenization to obtain a homo-, geneous dispersion.
Into a 3-litre flask provided witn a stirrer, in which ~1 2 litres of decahydronaphthalene and one gram of polyox~ethylene I sorbitan (Trade mark ~Twin #80) were placed, 100 ml of the ,,lhomogeneous dispersion of the mixture of Liquids C and A were introduced, and after one hour of stirring at 1000 rpm, the ~mixture was re-dispersed into 10 litres of ethanol to obtain an insoluble material. The insoluble material was collected by filtration and washed repeatedly with ethanol to remove decahydronaphthalene, and then, the washed material was again dispersed into one litre of an aqueous 2 % ammonium hydroxide solution. On neutralizing the dispersion, spherical particles were deposited. The particles were collected by filtration, and washed with water to obtain a composite material o~ the present invention (Specimen K) as spherical particles of 0.1 to 1.0 mm in diameter.
Results of examination of the surface of the particles of Specimen K as in Example 16 showed the adherence of platelets and blood corpuscles on the surface of the particles, Specimen X.
EXAMPLE 18:
After pouring the aqueous mixture of collagen fiber and de-N-acetylated chitin prepared in Example 6 onto a plate :¦~ of 200 mm square to a thickness of 5 mm and leaviny still for .
I one hour, the plate with the mixture was immersed into an ¦aqueous 0.lN ammonium hydroxide solution to neutralize the 1,1 ,1 ; .
'' " ` ''.
1 11475~7 1, .
Il acidity, and washed with de-ionized water to remove salts.
,~~ The thus obtained material was immersed into an aqueous 0.1 M
disodium phosphate solution containing 0.5 % by weight of form- ¦
aldehyde at 30C for one hour and then washed with water.
The thus obtained formed material was freeze-dried to be a sponge-form material. The sponge-form material Il showed an absorption of aqueous physiological saline solution ¦ of 20 g per g of the material and further, it shows a strong coagulation activity to bloods, and accordingly, it has been lo found that the material is possib~'ly applicable as a menstrual pad material and as a garrot tourniquet for urgent cases.
EXAMPLE l9:
~ The aqueous mixture of collagen fiber, gelatin and ¦I de-N-acetylated chitin of Example 2 was treated in the same manner as in Example 18 to obtain a spongy-form ma~erial. The thus obtained material showed an absorption ability to an aqueous physiological saline solution of 21 g/g material, and a strong ~`~ coagulating property to human blood. Accordingly, it was found that the material is applicable as the material for menstrual pads and tourniquets for urgent cases.
EXAMPLE 20:
Into a solution prepared by dissolving 1 g of glucose-~ isomerase(2000 U/g; manufactured by Nagase Sangyo Co. Ltd.) I into an aqueous O.lM disodium phosphate solution, 30 g of the spherical particles (Specimen ~ of Example 16) were added, and ; I after stirring the mixture for 2 hours at 5~C, 2 ml of an , .
' " ' ', ` ~1475~7 aqueous 25 % by weight glutaraldehyde solution was added to the mixture. After leaving the mixture as it is for 5 hours, solid materials were recovered by filtration, washed with an aqueous phosphoric buffer solution of pH of 7.0 and added to S00 ml of an aqueous phosphoric buffer solution containing 2 g of glucose. After thus treating the mixture for 60 min at 70-C, the contents of glucose and fructose of the solid material were 600 and 1400 mg, respectively, together with the fixed enzyme I (glucose-isomerase) of 70 ~ of the applied amount.
lo In addition, separately, the pH of the solution of collagen and de-~-acetylated chitin before re-dispersion in il Example 16 was adjusted to 6.0, and 1 g of glucose-isomerase ~ was added to the solution at 5-C. After re-dispersing the ¦I mixture as in Example 16, the solid matter was recovered by ¦ filtration and washed with ethanol and then with an aqueous phosphoric buffer solution of pH of 7Ø
It was found that 85 ~ of the enzyme (glucose-isomerase) applied was fixed onto the solid matter.
These results show that the thus obtained material is possibly applied as a carrier for fixed enzymes and microbial bodies.
EXAMPLE 21:
The same procedures as in Example 20 were carried out ; except for using Specimen X obtained in Example 17 instead of Specimen H used in Example 20. The amounts of glucose and ¦ fructose on Specimen K after the treatment at 70-C for 60 min '.~, , - 44 -, 1 ~1475()7 ., were 600 and 1400 mg, respectively with the fixed rate of glucose isomerase of 70 ~.
¦ From these results, it has been found that a compo-site material according to the present invention, i.e., Specimen K, is possibly applicable as a carrier of fixed enzymes and microbial bodies.
EXAMPLE 22: ~
Ater immersing Comparative Specimen A comprising a shaped material consisting only of collagen fibers into a lo homogeneous solution of a viscosity of 50 cP prepared by adding
5 g of de-N-acetylated chitin having a degree of de-N-acetylation of 85 % into one litre of an aqueous 1 ~ acetic acid solution at 20C for one hour, the excess aqueous solution was removed, and the thus treated material in a wet state was once immersed into an aqueous 0.1 N ammonia solution. Then the shaped material was washed with water to remove the attached ammonium acetate and ammonia to obtain a composite material according to the present invention.
The composite material showed the following mechanical properties of 395 kg/cm of tensile strength, 33 % of elonga-tion at break and 50 g cm/cm of tear strength. The immersed amount of de-N-acetylated chitin into collagen fibers was 2/100 by weight.
EXAMPLE 23:
In the same manner as in Example 22 except for using Comparativ Specimen B ootained in Example 2 and a solution !i il il - 45 -- .
-, ~
11~7507 (viscosity; 100 cP) of de-N-acetylated chltin having a degree of de-N-acetylation of 75 %, a composite material of the present invention was obtained. The mechanical properties thereof were: 400 kg/cm2 in tensile strength, 30 ~ in elongation at l break and 30 g cm/cm in tear strength. The content of de-N-¦l acetylated chitin of the composite material obtained in this example was 2 parts per 100 parts of collagen fiber.
'~
,
The composite material showed the following mechanical properties of 395 kg/cm of tensile strength, 33 % of elonga-tion at break and 50 g cm/cm of tear strength. The immersed amount of de-N-acetylated chitin into collagen fibers was 2/100 by weight.
EXAMPLE 23:
In the same manner as in Example 22 except for using Comparativ Specimen B ootained in Example 2 and a solution !i il il - 45 -- .
-, ~
11~7507 (viscosity; 100 cP) of de-N-acetylated chltin having a degree of de-N-acetylation of 75 %, a composite material of the present invention was obtained. The mechanical properties thereof were: 400 kg/cm2 in tensile strength, 30 ~ in elongation at l break and 30 g cm/cm in tear strength. The content of de-N-¦l acetylated chitin of the composite material obtained in this example was 2 parts per 100 parts of collagen fiber.
'~
,
Claims (14)
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A composite material comprising de-N-acetylated chitin having a degree of de-N-acetylation of 0.5 to 1.0, and fibrous collagen in a weight ratio of 0.01/99.99 to 99/1.
2. Composite material of Claim 1, wherein the fibrous collagen is partially replaced by gelatin and/or soluble collagen.
3. Composite material of Claim 1, wherein the fibrous collagen is previously cross-linked or acylated.
4. Composite material of any one of claims 1 to 3, which is further cross-linked.
5. A method for preparing a composite material of Claim 1, comprising bringing de-N-acetylated chitin into contact with fibrous collagen in an acidic medium followed by deacidify-ing the obtained product.
6. A method of Claim 5, wherein the contact of the de-N-acetylated chitin with the fibrous collagen is effected by mixing an aqueous acidic solution of the de-N-acetylated chitin and an aqueous dispersion of fibrous collagen.
7. A method of Claim 5, wherein the contact of the de-N-acetylated chitin with the fibrous collagen is effected by immersing a previously shaped material of the fibrous collagen into an aqueous acidic solution of the de-N-acetylated chitin.
8. A method of any one of Claims 5 to 7, wherein the fibrous collagen is partially replaced by gelatin and/or soluble collagen.
9. A method of Claim 5, wherein the fibrous collagen is previously cross-linked or acylated before the contact with the de-N-acetylated chitin.
10. A method of Claim 5, wherein the deacidifying step is effected by the addition of an aqueous alkaline solution the evaporating removal of the acid or the electrodeposition.
11. A method of Claim 5, wherein the resultant composite material is further cross-linked.
12. The composite material of any one of Claims 1, 2 or 3 in the form of an edible food additive.
13. The composite material of any one of Claims 1, 2 or 3 in the form of a base material for immobilizing enzyme.
14. The composite material of any one of Claims 1, 2 or 3 in the form of an edible casing for meats.
Applications Claiming Priority (4)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP3671280A JPS56133344A (en) | 1980-03-21 | 1980-03-21 | Novel composite material |
| JP36711/80 | 1980-03-21 | ||
| JP3671180A JPS56131639A (en) | 1980-03-21 | 1980-03-21 | Protein-chitosan composition |
| JP36712/80 | 1980-03-21 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1147507A true CA1147507A (en) | 1983-06-07 |
Family
ID=26375794
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000373335A Expired CA1147507A (en) | 1980-03-21 | 1981-03-18 | Composite material of de-n-acetylated chitin and fibrous collagen |
Country Status (5)
| Country | Link |
|---|---|
| US (1) | US4378017A (en) |
| EP (1) | EP0038628B1 (en) |
| AU (1) | AU535551B2 (en) |
| CA (1) | CA1147507A (en) |
| DE (1) | DE3171293D1 (en) |
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| IT1144925B (en) * | 1981-10-08 | 1986-10-29 | Anic Spa | CHITOSAN QUARRY FIBERS AND PROCEDURE FOR THEIR OBTAINING |
| FR2517315B1 (en) * | 1981-11-30 | 1985-12-20 | Tech Cuir Centre | PROCESS FOR THE PREPARATION OF NEW FORMS OF COLLAGEN, NATIVE OR DERETICULATED, WITH PRESERVED HELICOIDAL STRUCTURE, ASSOCIATED WITH MUCOPOLYSACCHARIDES AND THEIR APPLICATIONS IN PARTICULAR IN THE COSMETOLOGICAL, PHARMACEUTICAL, ANALYTICAL AND OTHER FIELDS |
| CA1213520A (en) * | 1982-03-17 | 1986-11-04 | Abe Widra | Hydrophilic biopolymeric copolyelectrolytes, and biodegradable dressings comprising same |
| US4570629A (en) * | 1982-03-17 | 1986-02-18 | University Of Illinois Foundation | Hydrophilic biopolymeric copolyelectrolytes, and biodegradable wound dressing comprising same |
| AU569112B2 (en) * | 1983-02-08 | 1988-01-21 | Nitta Gelatin Co. Ltd. | Crosslinked collagen products |
| US5290752A (en) * | 1984-03-16 | 1994-03-01 | Unitika Ltd. | Method for preparation of a shaped chitin body containing a physiologically active substance |
| US4664658A (en) * | 1984-11-08 | 1987-05-12 | Mitsubishi Monsanto Chemical Company | Medical material and process for its production |
| US4861714A (en) * | 1985-04-04 | 1989-08-29 | Verax Corporation | Weighted collagen microsponge for immobilizing bioactive material |
| JPS61240963A (en) * | 1985-04-18 | 1986-10-27 | ユニチカ株式会社 | Wound covering protective material |
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| US5100783A (en) * | 1985-05-10 | 1992-03-31 | Verax Corporation | Weighted microsponge for immobilizing bioactive material |
| CA1292597C (en) * | 1985-12-24 | 1991-12-03 | Koichi Okita | Tubular prothesis having a composite structure |
| FR2616318A1 (en) * | 1987-06-15 | 1988-12-16 | Centre Nat Rech Scient | ARTIFICIAL SKIN AND PROCESS FOR PREPARING THE SAME |
| EP0318286A3 (en) * | 1987-11-26 | 1990-10-31 | Katakura Chikkarin Co., Ltd. | A substratum for cell culture and its production and use |
| US5605835A (en) * | 1988-05-23 | 1997-02-25 | Regents Of The University Of Minnesota | Bioreactor device with application as a bioartificial liver |
| US5510418A (en) * | 1988-11-21 | 1996-04-23 | Collagen Corporation | Glycosaminoglycan-synthetic polymer conjugates |
| US5395620A (en) * | 1989-01-31 | 1995-03-07 | Coletica | Biodegradable microcapsules having walls composed of crosslinked atelocollagen and polyholoside |
| NZ244737A (en) * | 1989-02-21 | 1993-09-27 | Viskase Corp | Food packaging polymeric film containing antibiotic material; method of treating foodstuff and food casing therefor |
| US5100429A (en) * | 1989-04-28 | 1992-03-31 | C. R. Bard, Inc. | Endovascular stent and delivery system |
| US6004261A (en) * | 1989-04-28 | 1999-12-21 | C. R. Bard, Inc. | Formed-in-place endovascular stent and delivery system |
| US4990158A (en) * | 1989-05-10 | 1991-02-05 | United States Surgical Corporation | Synthetic semiabsorbable tubular prosthesis |
| US5217495A (en) * | 1989-05-10 | 1993-06-08 | United States Surgical Corporation | Synthetic semiabsorbable composite yarn |
| US5376118A (en) * | 1989-05-10 | 1994-12-27 | United States Surgical Corporation | Support material for cell impregnation |
| US5147400A (en) * | 1989-05-10 | 1992-09-15 | United States Surgical Corporation | Connective tissue prosthesis |
| US5093319A (en) * | 1989-10-31 | 1992-03-03 | Pfizer Hospital Products Group, Inc. | Use of derivatives of chitin soluble in aqueous solutions for preventing adhesions |
| US4994277A (en) * | 1989-10-31 | 1991-02-19 | Pfizer Hospital Products Group, Inc. | Use of xanthan gum for preventing adhesions |
| US5092841A (en) * | 1990-05-17 | 1992-03-03 | Wayne State University | Method for treating an arterial wall injured during angioplasty |
| JP2582979B2 (en) * | 1990-10-09 | 1997-02-19 | ファイザー インク | Quaternary ammonium salts derived from chitosan |
| FR2669629A1 (en) * | 1990-11-22 | 1992-05-29 | Morelle Jean | New inhibitors of proteolytic and decarboxylase enzymes |
| DE4107239A1 (en) * | 1991-03-07 | 1992-09-10 | Hoechst Ag | TUBULAR FOOD COVER WITH IMPROVED REELABILITY |
| DE4121068A1 (en) * | 1991-03-07 | 1992-09-10 | Hoechst Ag | TUBULAR FOOD COVER WITH IMPROVED REELABILITY |
| DE4127604A1 (en) * | 1991-08-21 | 1993-02-25 | Hoechst Ag | TUBULAR FOOD COVER WITH IMPROVED ADHESIVE PROPERTIES OVER THE FILLING MATERIAL |
| US5846952A (en) * | 1993-12-01 | 1998-12-08 | Marine Polymer Technologies, Inc. | Methods and compositions for poly-β-1-4-N-acetylglucosamine drug delivery |
| US6063911A (en) * | 1993-12-01 | 2000-05-16 | Marine Polymer Technologies, Inc. | Methods and compositions for treatment of cell proliferative disorders |
| US5858350A (en) * | 1993-12-01 | 1999-01-12 | Marine Polymer Technologies | Methods and compositions for poly-β-1→4-N-acetylglucosamine cell therapy system |
| US6743783B1 (en) * | 1993-12-01 | 2004-06-01 | Marine Polymer Technologies, Inc. | Pharmaceutical compositions comprising poly-β-1→4-N-acetylglucosamine |
| US5635493A (en) * | 1993-12-01 | 1997-06-03 | Marine Polymer Technologies, Inc. | Methods and compositions for poly-β-1-4-N-acetylglucosamine chemotherapeutics |
| US5686115A (en) * | 1993-12-01 | 1997-11-11 | Marine Polymer Technologies, Inc. | Poly-β-1→4-N-acetylucosamine copolymer composition with collagen |
| US5624679A (en) * | 1993-12-01 | 1997-04-29 | Marine Polymer Technologies, Inc. | Methods and compositions for poly-β-1-4-N-acetylglucosamine biological barriers |
| US5622834A (en) * | 1993-12-01 | 1997-04-22 | Marine Polymer Technologies, Inc. | Method of isolating poly-β-1-4-N-acetylglucosamine from microalgal culture |
| ATE310839T1 (en) * | 1994-04-29 | 2005-12-15 | Scimed Life Systems Inc | STENT WITH COLLAGEN |
| SE9402531L (en) * | 1994-07-19 | 1996-01-20 | Medicarb Ab | wound healing agent |
| US5931165A (en) * | 1994-09-06 | 1999-08-03 | Fusion Medical Technologies, Inc. | Films having improved characteristics and methods for their preparation and use |
| US5622740A (en) * | 1994-11-14 | 1997-04-22 | Devro Limited | Method for preparing edible casings and compositions thereof |
| US5603884A (en) * | 1994-11-18 | 1997-02-18 | Viskase Corporation | Reinforced cellulosic film |
| CN1065773C (en) * | 1994-12-28 | 2001-05-16 | 天津纺织工学院膜天膜技术工程公司 | Flexible suture of absobable biological material for medical use |
| US6150581A (en) * | 1995-06-07 | 2000-11-21 | United States Surgical Corporation | Chitosan/alginate anti-adhesion barrier |
| JPH09122227A (en) * | 1995-10-31 | 1997-05-13 | Bio Eng Lab:Kk | Medical material and manufacture thereof |
| NL1018871C2 (en) * | 2001-09-03 | 2003-03-05 | Ktc Beheer B V | Process for preparing an extruded food product. |
| RU2240830C1 (en) * | 2003-12-26 | 2004-11-27 | ФГУП Государственный научно-исследовательский институт особо чистых биопрепаратов | Wound coating and method for its preparing |
| NZ550964A (en) | 2004-04-28 | 2011-05-27 | Angiodevice Internat Gmbh | Compositions and systems for forming crosslinked biomaterials and associated methods of preparation and use |
| CZ300697B6 (en) * | 2006-04-11 | 2009-07-22 | Výzkumný ústav potravinárský Praha, v.v.i. | Functional symbiotic edible food packages |
| ES2552842T3 (en) * | 2007-02-19 | 2015-12-02 | Marine Polymer Technologies, Inc. | Hemostatic compositions and therapeutic regimens |
| PL2223941T3 (en) * | 2009-02-27 | 2019-04-30 | Rohm & Haas | Rapid cure carbohydrate composition |
| EP2467172A1 (en) * | 2009-08-10 | 2012-06-27 | Osseous Technologies Of America | Self-supporting collagen tunnel for guided tissue regeneration and method of using same |
| WO2011127591A1 (en) * | 2010-04-15 | 2011-10-20 | Covalon Technologies Inc. | Reinforced tissue shields |
| AU2011239466B2 (en) | 2010-04-15 | 2015-01-22 | Marine Polymer Technologies, Inc. | Anti-bacterial applications of poly -N-acetylglucosamine nanofibers |
| EP3501284A1 (en) | 2011-04-15 | 2019-06-26 | Marine Polymer Technologies, Inc. | Treatment of skin deseases with poly-n-acetyl glucosamine nanofibers |
| FR3016882A1 (en) | 2014-01-30 | 2015-07-31 | Sofradim Production | PROCESS FOR THE PREPARATION OF HIGH-DEGREE ACETYLATION CHITOSAN |
| US20180325128A1 (en) * | 2017-05-09 | 2018-11-15 | Nitta Casings Inc. | Collagen casings with added chitosan |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB1040770A (en) * | 1963-11-20 | 1966-09-01 | Unilever Ltd | Foodstuffs |
| GB1109509A (en) * | 1964-09-08 | 1968-04-10 | Unilever Ltd | Treatment of proteins particularly sausage casings |
| US3833744A (en) * | 1972-08-24 | 1974-09-03 | Gen Mills Inc | Texturizing materials for foods |
| GB1515963A (en) | 1975-07-15 | 1978-06-28 | Massachusetts Inst Technology | Crosslinked collagen-mucopolysaccharide composite materials |
| US4280954A (en) * | 1975-07-15 | 1981-07-28 | Massachusetts Institute Of Technology | Crosslinked collagen-mucopolysaccharide composite materials |
| JPS5431458A (en) * | 1977-08-15 | 1979-03-08 | Agency Of Ind Science & Technol | Molded resin article containing chitin and its manufacture |
| US4264155A (en) * | 1979-07-09 | 1981-04-28 | Opticol Corporation | Collagen contact lens |
| US4301067A (en) * | 1979-06-05 | 1981-11-17 | Kureha Kagaku Kogyo Kabushiki Kaisha | Chitin containing poly-ion complex |
| JPS55165999A (en) * | 1979-06-12 | 1980-12-24 | Kureha Chemical Ind Co Ltd | Manufacture of collagen fiber article |
| US4260228A (en) * | 1980-01-21 | 1981-04-07 | Opticol Corporation | Collagen gel contact lens and method of preparation |
-
1981
- 1981-03-11 US US06/242,419 patent/US4378017A/en not_active Expired - Lifetime
- 1981-03-18 CA CA000373335A patent/CA1147507A/en not_active Expired
- 1981-03-18 AU AU68521/81A patent/AU535551B2/en not_active Ceased
- 1981-03-20 DE DE8181301212T patent/DE3171293D1/en not_active Expired
- 1981-03-20 EP EP81301212A patent/EP0038628B1/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| AU535551B2 (en) | 1984-03-29 |
| EP0038628A2 (en) | 1981-10-28 |
| EP0038628A3 (en) | 1982-06-09 |
| US4378017A (en) | 1983-03-29 |
| EP0038628B1 (en) | 1985-07-10 |
| DE3171293D1 (en) | 1985-08-14 |
| AU6852181A (en) | 1981-09-24 |
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