JPS6159737B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a wound treatment material used for treating wounds such as cuts, abrasions, burns, surgical wounds, ulcers on the body surface, tooth extraction sockets, etc. More specifically, the present invention relates to a wound protection material that can effectively promote the production of stabilized fibrin in a wound for a long period of time.

Gelatin sponge containing trompine is known as a therapeutic material applied to wounds such as cuts and abrasions, surgical wounds, tooth extraction sockets, etc. (U.S. Pat. No. 2,558,395, Japanese Patent Publication No. 31-4644). . Thrombin contained in gelatin acts on fibrinogen in the wound area to form a fibrin clot to stop bleeding, but the fibrin produced by thrombin is called unstabilized fibrin and is susceptible to acids, urea, etc. Because it is soluble and susceptible to plasmin degradation, wound healing is often significantly delayed.

The present inventors have conducted intensive studies to obtain a wound treatment material that can effectively promote the production of stabilized fibrin on the wound surface for a long period of time, and have found that thrombin and blood coagulation factors (hereinafter abbreviated as factors) have been developed. The present invention has been achieved by discovering that by immobilizing both fibrinogen, stabilized fibrin can be produced via non-stabilized fibrin.

That is, the present invention provides a wound treatment material comprising a structure in the form of a monofilament, a fiber aggregate, a film, a sponge, etc., in which thrombin and factors are immobilized on the structure. The present invention is a wound treatment material that can effectively promote the production of stabilized fibrin in a wound.

The wound treatment material in the present invention refers to a known treatment material consisting of a structure having a shape such as a fiber aggregate such as monofilament, cotton, paper, nonwoven fabric, woven fabric, or knitted fabric, film, or sponge. For example, surgical sutures have the form of monofilaments, twisted threads, knitted threads, and the like. Absorbent pads, bandages, burn dressings, tooth extraction socket fillings, etc. used to protect wounds and facilitate their healing have the form of cotton, paper, non-woven fabrics, woven fabrics, knitted fabrics, films, sponges, etc.

Materials constituting these structures include, for example, cellulose such as cotton, hemp, and pulp, regenerated cellulose obtained by the viscose method and cuprammonium method, cellulose derivatives such as cellulose acetate, agarose, textolan, pullulan starch, and argyl. acids, polysaccharides such as pectic acid,
Proteins such as silk, collagen, gelatin casein, and wool; synthetic polyamino acids such as polyglutamic acid, polyalanine, polyphenylalanine, and polylysine; polyesters such as polyethylene terephthalate, polyglycolic acid, polylactic acid, and polybutylene terephthalate; nylon 6 and nylon 11. ,
Nylon 12, Nylon 6,6, Nylon 6,10,
Polyamides such as poly(m-phenylene isophthalamide) and poly(p-phenylene terephthalamide), olefins such as polyethylene, polypropylene, polybutadiene, and polyisoprene, polymers of dienes, polyvinyl chloride, polyvinylidene chloride, chlorine Chlorinated polyolefins such as polyethylene, polymers of N-vinyl compounds such as N-vinylpyrrolidone and N-vinylamine, styrene,
Polymers of aromatic vinyl compounds such as a-methylstyrene, vinylpyridine, divinylstyrene, polyvinyl alcohol and its ester derivatives such as polyvinyl acetate, polymers of unsaturated aldehydes such as polyacrolein, acrylic acid, methacrylic acid, fumaric acid , polymers of unsaturated carboxylic acids such as maleic acid, polymers of unsaturated carboxylic acid esters such as methyl acrylate, ethyl acrylate, methyl methacrylate, and ethyl methacrylate, acrylic anhydride, methacrylic anhydride,
Polymers of unsaturated carboxylic acid anhydrides such as maleic anhydride, polymers of unsaturated nitriles such as acrylonitrile and memethacroylnitrile, polymers of unsaturated carboxylic acid amides such as acrylamide and methacroylamide, and polymers such as polyethyleneimine. Polyethers such as alkylene imine, polyethylene oxide, polypropylene oxide, polymethylene oxide, and polyphenylene oxide, silicone resins such as dimethylpolysiloxane and methylphenylpolysiloxane, and polyisocyanates such as toluene diisocyanate and xylene diisocyanate. Examples include polymeric substances such as polyurethane derived from polyester, polyethylene glycol, polypropylene glycol, polyester having hydroxyl groups at both ends, and rubber with added combustion. Furthermore, linear copolymers, crosslinked copolymers, graft copolymers, and block copolymers containing monomer components of the above-mentioned polymeric substances can also be used as materials constituting the structure. The appropriate molecular weight range for the polymer substance is 7000 to have the necessary mechanical strength as a material for wound treatment.
or more, preferably 10,000 or more.

In the present invention, it is necessary that both thrombin and a factor be immobilized on the above-mentioned structure. Immobilization of thrombin alone produces non-stabilized fibrin, but the non-stabilized fibrin cannot be converted into stabilized fibrin. On the other hand, fibrin is not produced when only the factor is immobilized. However, by immobilizing both thrombin and factors as in the present invention, stabilized fibrin can be produced from fibrinogen via non-stabilized fibrin.

Thrombin used in the present invention is a proteolytic enzyme that can convert fibrinogen to fibrin. Thrombin is isolated from blood of humans, cows, pigs, etc., but when applied to human wounds, it is preferable to use human thrombin.

The factors used in the present invention are called fibrin stabilizing factors, and act directly on non-stabilized fibrin.
It is a factor involved in the generation of isobeptide bonds between fibrin molecules. The factor is isolated from blood or placenta of humans, cows, etc., but when applied to human wounds, it is preferable to use a factor derived from humans.

Thrombin and factors can be immobilized by binding or adsorption to a structure having a shape such as a monofilament, a fiber aggregate film, or a sponge. The binding of thrombin and factors to structures constituting wound treatment materials has been described, for example, by O. Zaborsky,
Conventionally known covalent bonding methods and ionic bonding methods such as those described in “Immobilized Enzymes” CRC Press, 1973 can be employed, and physical adsorption methods and entrapment methods can also be used for adsorption. Can be adopted.

The structures constituting the wound treatment material described above include carboxyl groups, chlorotriazinyl groups, azide groups, diazonium groups, epoxy groups, formyl groups,
Reactive functional groups capable of forming covalent bonds such as acid anhydride groups, bromoacetyl groups, isocyanate groups, iminocarbonate groups, or amino groups, ammonium groups, carboxyl groups, carboxylate groups,
When the structure has an ion exchange group such as a sulfoxyl group or a sulfonate group, thrombin and the factor can be covalently bonded or ionically bonded by treating the structure with a solution containing thrombin and the factor. In addition, if the structure does not have any or only a few functional groups and ion exchange groups that can form covalent bonds, these groups may be introduced into the structure by a polymer reaction, and then thrombin and factors may be added to the structure. can be bonded to the structure.

Examples of binding thrombin and factors to various structures are listed below.

(1) Proteins such as silk, collagen, casein, and gelatin have amino groups and carboxyl groups, which are functional groups that can form covalent bonds and are also ion exchange groups. In some cases, thrombin and factors can be covalently or ionically bound without pretreatment. In the case of covalent bonding, dicyclohexylcarbodiimide, 1-cyclohexyl-3-(2-morpholininoethyl)-carbodiimide-meth-p-
Preferably, a dehydration condensation agent such as toluene sulfonate is used.

Furthermore, by reacting a protein-based structure with a dialdehyde starch maleic anhydride copolymer or the like, formyl groups, acid anhydride groups, etc. can be introduced, and thrombin and factors can then be covalently bonded.

(2) In the case of structures based on cellulose or cellulose derivatives such as cotton, regenerated cellulose, cellulose acetate, or polyvinyl alcohol, these structures may be treated with polyaldehydes such as glutaraldehyde or dialdehyde starch. Thrombin and the factor can be covalently bonded by introducing a formyl group into the structure and then treating the structure into which the formyl group has been introduced with thrombin and the factor. Instead of borealdehyde, bromocyanide can be used to create iminocarbonate groups, bromoacetyl groups, polymaleic anhydride groups can be used to create acid anhydride groups such as maleic anhydride copolymers, and polyisocyanates can be used to create isocyanate groups such as toluene diisocyanate and hexamethylene diisocyanate. Epoxy groups can be introduced into the structure using nerto groups, polyglycidine derivatives such as ethylene glycol, tetramethylene glycol, and glycerin, and chlortriazinyl groups can be introduced into structures using cyanuric chloride. It can also be used to bind thrombin and factors. Structures into which formyl groups, iminocarbonate groups, bromoacetyl groups, acid anhydride groups, isocyanate groups, epoxy groups, and chlortriazinyl groups have been introduced are aminated by treatment with polyamines such as polyethyleneimine, and glycine, etc. is carboxylated by treatment with an aminocarboxylic acid. Thrombin and factors can be ionically bonded to the structure into which ion exchange groups such as amino groups and carboxyl groups have been introduced in this way.

(3) In the case of structures made from polymeric substances with terminal carboxyl groups, such as polyamides and polyesters, these structures are treated with polyethyleneimine in the presence of a dehydration condensation agent, and then treated with maleic anhydride copolymer. Acid anhydride groups are introduced into the structure by treatment with coalescence. By treating the structure into which an acid anhydride group has been introduced with thrombin and a factor, thrombin and the factor can be covalently bonded. When a structure into which an acid anhydride group has been introduced is hydrolyzed, a carboxyl group, which is an ion exchange group, is generated. Thrombin and factors can be ionically bonded to the structure into which carboxyl groups have been introduced in this way.

In addition, when manufacturing the wound treatment material of the present invention, thrombin and factors may be physically adsorbed or encased in structures such as monofilaments, fiber aggregates, films, and sponges as follows. It can be adsorbed by methods such as methods. That is, thrombin and the factor can be physically adsorbed by dissolving the thrombin and the factor in a solvent that can wet the structure and treating the structure with this solution. The entrapment method is a method in which thrombin and factors are encapsulated in a fine lattice of gel so that they cannot be released. Suitable materials for use in this method include absorbable substances such as collagen, gelatin, polyglycolic acid, polylactic acid, glycolic acid-lactic acid copolymer, polyglutamic acid, and amylose.

In order to produce the wound treatment material of the present invention, in addition to the method of binding or adsorbing thrombin and factors to a structure having a shape such as a monofilament, fiber aggregate, film, or sponge as described above, It is also possible to manufacture a structure by first binding or adsorbing thrombin and factors to the material itself before processing it into a structure, and then processing the material to which thrombin and factors have been bound or adsorbed into a structure. For example, structures such as monofilaments, cotton, paper, nonwoven fabrics, woven fabrics, knitted fabrics, films, sponges, etc. are obtained by using a polymer substance to which thrombin is bound or adsorbed in advance, and the wound treatment material of the present invention is obtained. can be manufactured.

When immobilizing thrombin and factors by any of the above methods, thrombin and factors may be immobilized at the same time, or thrombin may be immobilized first and then the factors may be immobilized, The opposite may be true.

When manufacturing the wound treatment material of the present invention,
Calcium ions involved in factor activation can be immobilized. Furthermore, when manufacturing the wound treatment material of the present invention, pharmaceuticals such as antiplasmin agents, bactericides, antibiotics, hormones, etc. can be immobilized on the structure along with thrombin and factors, if necessary. Antiplasmin is an inhibitor of the fibrinolytic enzyme plasmin, and thus suppresses fibrinolytic activity (ie, fibrinolytic activity) by inhibiting plasmin. Therefore, a wound treatment material in which antiplasmin is immobilized together with thrombin and factors can promote fibrin production by suppressing fibrinolytic activity. Examples of antiplasmin include aprotinin extracted from bovine lungs, bepstatin isolated from microbial culture fluid, leupepsin,
Natural substances such as antivaine and chymostatin, ε
-aminocaproic acid, tranexamic acid, and gabexate mesylate, among which ε-aminocaproic acid and traneximic acid are particularly preferably used.

The wound protection material of the present invention can be used to protect wounds such as cuts and abrasions, burn surfaces, remains on the body surface, tooth extraction sockets, etc., and can effectively stabilize fibrin in the wound for a long period of time. Since it can promote production, it has the effect of significantly speeding up treatment and relieving local pain at the same time. In addition, collagen is used as a raw material for wound protection materials.
When absorbable substances such as gelatin, polyglycolic acid, polylactic acid, glycolic acid-lactic acid copolymer, polyglutamic acid, and amylose are used, there is no need to remove them after the purpose is achieved, so they can be absorbed into the body, especially at anastomotic sites. It is preferably used in areas where promotion of stabilized fibrin formation is required, with gelatin, polyglycolic acid, and polylactic acid being particularly preferred.

The wound protection material of the present invention can be easily sterilized using a gas sterilizer such as ethylene oxide. Furthermore, sterilization can also be carried out by methods such as irradiation with X-rays, gamma rays, neutrons, electrons, and the like.

Next, the present invention will be explained in more detail with reference to Examples. As the thrombin, human plasma thrombin from Midori Juji Co., Ltd. was used, and as the factor, a concentrated dry preparation of placenta-derived factors from Behringwerke was used. The factor preparation has a factor activity equivalent to 250 ml of fresh human plasma per bottle.

Further, the production of stabilized fibrin was confirmed as follows. That is, after treating a structure with immobilized thrombin and factors with an aqueous solution containing Ca ⁺⁺ to activate the factors,
The structure was placed in a 0.5% fibrinogen aqueous solution and left at 37°C for 10 minutes, and then it was confirmed whether the fibrin produced on the surface of the structure was stabilized fibrin. Non-stabilized fibrin dissolves in 1% monochloroacetic acid, while stabilized fibrin dissolves in 1% monochloroacetic acid.
% monochloroacetic acid, this was confirmed by examining the solubility of the fibrin membrane formed as described above in 1% monochloroacetic acid.

Example 1 Sharp et al.'s method [AKSharp, G. Kay, MD
Lilly, Biotechnology and Bioengineering, 11
Vol., p. 363 (1969)], surgical gauze (5 cm x 5 cm) was coated with cyanuric chloride, followed by N-
After reacting with (3-aminopropyl)-diethanolamine, 4 ml of factor aqueous solution (1 bottle of concentrated dry preparation of factor dissolved in 4 ml of water) and 4 ml of thrombin saline solution (100 units/ml)
It was immersed in a mixed solution consisting of and reacted at 25°C for 10 minutes. After adding an excessive amount of N-(3-aminopropyl)-diethanolamine aqueous solution to react the remaining active chlorine, it was washed with physiological saline.
The fibrin formed on the surface of the surgical gauze on which thrombin and factors were immobilized was not dissolved by 1% monochloroacetic acid.

For comparison, immobilization was carried out in the same manner as above except that only a thrombin solution or only a factor solution was used instead of the mixture of thrombin and factor. As a result, fibrin formed on the surface where only thrombin was immobilized was dissolved by 11% monochloroacetic acid.
On the other hand, no fibrin was formed on the surface where only the factor was immobilized.

Example 2 Taffeta made of polyethylene terephthalate (10cm
× 10 cm) in a mixed solution consisting of a 10% polyethyleneimine aqueous solution and 5 times the amount of methanol,
It was left at room temperature for 30 minutes. Next, a methanol solution of 4% by weight dicyclohexylcarbodiimide was added in an amount twice the volume of the aqueous polyethyleneimine solution, and the mixture was further left at room temperature for 2 hours. After soaking, the taffeta was washed with methanol and then dried. The polyethylene-treated taffeta thus obtained was then placed in an acetone solution containing 3% by weight of methyl bisether-maleic anhydride copolymer, allowed to stand at room temperature for 2 hours, and then washed with acetone. After drying the obtained taffeta, it was left in an aqueous solution containing thrombin and factors for 24 hours at 4°C in the same manner as in Example 1, and then washed with physiological saline. The fibrin formed on the surface of the taffeta on which thrombin and factors were immobilized thus obtained was not dissolved by 1% monochloroacetic acid.

Example 3 Gelatin sponge manufactured by Yamanouchi Pharmaceutical Co., Ltd. (5
cm x 2.5 cm x 0.5 cm) was immersed in 10 ml of the aqueous solution containing thrombin and factor used in Example 1 for 10 minutes, and then dried for 20 hours. The fibrin formed on the surface of the obtained gelatin sponge was not dissolved in 1% monochloroacetic acid.

Example 4 Commercially available polyglycolic acid knitted suture thread (American Cyanamid Company) and Special Publication 1974-
A polylactic acid knitted suture thread prepared with reference to Japanese Patent No. 31696 was treated in the same manner as in Example 3. As a result, the fibrin formed on the surface of any of the sutures on which thrombin and factors were immobilized was not dissolved by 1% monochloroacetic acid.

Claims (1)

[Claims] 1. Monofilament, fiber aggregate, film,
A wound treatment material consisting of a structure in the shape of a sponge, characterized in that thrombin and blood coagulation factors are immobilized on the structure, which effectively promotes the production of stabilized fibrin in the wound for a long period of time. material for wound treatment. 2. The wound treatment material according to claim 1, wherein the structure is made of an absorbent substance. 3. The wound treatment material according to claim 2, wherein the absorbent substance is gelatin. 4. The wound treatment material according to claim 2, wherein the absorbent substance is polyglycolic acid. 5. The wound treatment material according to claim 2, wherein the absorbent substance is polylactic acid.