JPH0663117A - Chitin composition - Google Patents

Abstract

PURPOSE:To obtain a chitin composition which has biodegradability, good affinity with a bone forming factor, excellent sustained release property for a bone forming factor and has no foreign matter reaction in a living body and further has an effect of assisting the bone forming reaction positively by incorporating a bone forming factor into a chitin. CONSTITUTION:A bone forming factor is contained in a chitin to form a chitin composition. As a manufacturing method for a chitin composition containing a bone forming factor, for example, a bone forming factor is suspended in a solvent, chitin is dipped in the suspension, and then dried. Any solvent which does not deactivate a bone forming factor may be used. At the time of forming a chitin compacted article, a bone forming factor may be mixed therein. The chitin compact containing the thus obtained bone forming factor can be used as it is, and can be powdered again to be used. The chitin composition is excellent in affinity with a living body, has no foreign matter reaction in the living body, has good sustained release property for a bone forming factor and exhibits bone forming performance sufficiently.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a chitin composition, and more particularly to chitin which controls the release of bone morphogenetic protein in vivo to induce bone formation and is decomposed in vivo together with bone formation. It relates to a composition.

[0002]

2. Description of the Related Art Conventionally, in the fields of orthopedics and oral surgery, autologous bone grafting has been performed as a method for prosthesis of a bone defect portion in a living body caused by trauma, excision or the like. Autologous bone graft has better engraftment to the transplant bed than performing allogeneic bone graft or xenogeneic bone graft, but there is a limit to the amount of bone that can be collected, and the formation of new surgical wounds for acquiring bone graft. There are drawbacks such as the risk of infection due to infection and prolonged distress to the patient. As an alternative to the autologous bone transplantation, a method of using a metal such as stainless steel or a titanium alloy as an artificial biomaterial has been performed because of the remarkable development of biomaterial and the easy availability. However, in the method using these artificial biomaterials, the material strength is excellent, but the affinity with living tissues is poor.

As a method for improving this point, surface treatment with a biocompatible material such as coating of the surface of such a material with hydroxyapatite or the like has been carried out to improve the affinity with the surrounding tissue, but it has not been improved yet. Not enough.

On the other hand, in recent years, biodegradable polymers such as polylactic acid, polylactic acid-polyglycolic acid copolymer, polyglycolic acid, poly-β-hydroxybutyrate and poly-ε-caprolactone have been used as biocompatible materials. Many studies have been conducted on polymer materials such as aliphatic polyesters and copolymers thereof with hydroxyaromatic carboxylic acids, or materials obtained by polymerizing these polymer materials with hydroxyapatite and tricalcium phosphate. However, when hydrolyzed in vivo, these materials have reduced mechanical strength and fatigue deterioration, and show almost no effect on bone formation, and merely damage the tissue in terms of biocompatibility. It is just a material without.

Under these circumstances, as bone-forming materials, physiologically active substances that differentiate and proliferate osteogenic cells and chondrocytes isolated from mouse Dunn osteosarcoma and human osteosarcoma, human bones, bovine bones, genes A bone-shaped material made of a complex of collagen and a bone morphogenetic factor obtained by recombination has been proposed. (JP-A-60-253455, JP-A-62-89629
However, when collagen is used, since the collagen is a material derived from a heterogeneous solid (cattle, pig, etc.), its molecular weight, amino acid composition, water retention amount, etc. are not constant, and the telopeptide part having antigenicity is used. Since it is difficult to completely remove the bone marrow, a foreign body reaction occurs in the body, and bone morphogenetic factors are phagocytosed by foreign body giant cells and other phagocytes,
The bone forming ability is not sufficiently expressed.

[0006] Further, there is known a medical material in which a drug is contained in a material such as polylactic acid or polyglycolic acid instead of collagen. However, according to this polylactic acid and the like, it is necessary to perform heat melting and heat formation when mixing drugs (Japanese Patent Publication No. 17525/50, Japanese Patent Publication No. 181029/1985).
When combined with bone morphogenetic factors, heat-induced degeneration,
It cannot be used because it deteriorates. Furthermore, as a method of mixing a drug using polylactic acid, polyglycolic acid or the like without heating and melting, polylactic acid or the like is dissolved using a solvent such as methylene chloride or chloroform, and the drug is mixed with this. Although a method is known, in this method as well, not only the bone morphogenetic factor is decomposed by the solvent, but since the base material after the solvent is volatilized becomes porous, the bone morphogenetic factor is mostly released in the initial stage, There is almost no sustained release.

In order to solve the above problems, use of a base material obtained by reacting a polymer or copolymer of lactic acid and / or glycolic acid with polyethylene glycol as a support for an osteogenic factor. Has been proposed (Japanese Patent Laid-Open No. 1-23052). It has a gel-like property and is easily mixed with an osteogenic factor, but the reaction product of a polymer or copolymer of lactic acid and / or glycolic acid and polyethylene glycol is merely a carrier, and itself. Does not actively support the bone formation reaction.

As described above, the bone forming material has various problems, is biodegradable, has a good affinity with the bone forming factor, is excellent in the bone forming ability, and does not adversely affect the living body. Furthermore, the material which positively supports the bone formation reaction has not been found yet.

[0009]

The present invention is biodegradable, has a good affinity for bone morphogenetic factors, is excellent in sustained release of bone morphogenetic factors, and is more active without foreign body reaction in the living body. Another object of the present invention is to provide a chitin composition having a function of assisting an osteogenic reaction.

[0010]

Means for Solving the Problems The inventors of the present invention have conducted extensive studies to solve the above problems, and as a result, when chitin was used as a support, an excellent bone forming biomaterial that avoids the above problems was obtained. The present invention has been found out and the present invention has been achieved. That is, the gist of the present invention is a chitin composition characterized by containing an osteogenic factor in chitin.

The present invention will be described in detail below. The chitin used in the present invention is originally poly (N-acetyl-D-glucosamine) obtained by treating the exoskeleton of crustaceans and insects with hydrochloric acid and caustic soda to remove ash and protein substances. However, the chitin referred to in the present invention includes poly N-acetyl-D-glucosamine and its deacetylated product and chitosan, and further a salt formed with chitin and acids,
For example, acetate salt, hydrochloride salt, nitrate salt, phosphate salt, etc., and further the --OH group or --CH ₂ OH group of the glucosamine residue is esterified, etherified, carboxymethylated, hydroxymethylated or O-ethylated. Chitin derivatives are also included.

Deacetylation of chitin in the case of using the deacetylated chitin in the present invention can be carried out by a known method of treating chitin with an alkali. On this occasion,
The degree of deacetylation can be easily adjusted by appropriately changing the alkali concentration used, the treatment temperature, the treatment time, and the like.

The term "deacetylation degree" as used herein means a value measured by the following method. About 2 g of the sample is put into 200 ml of a 2N-hydrochloric acid aqueous solution and stirred at room temperature for 30 minutes. Then, after filtering with a glass filter to remove the aqueous hydrochloric acid solution, the mixture was poured into 200 ml of methanol and mixed with 3 ml of water.
Stir for 0 minutes. This product is further filtered through a glass filter, put into 200 ml of fresh methanol, and stirred for 30 minutes. After repeating this washing operation with methanol four times, it was air-dried and vacuum-dried, and about 0.2 g thereof was precisely weighed, 100 ml was placed in an Erlenmeyer flask, 40 ml of ion-exchanged water was added, and the mixture was stirred for 30 minutes.
Then, this solution is subjected to neutralization titration with 0.1N-caustic soda aqueous solution using phenolphthalein as an indicator. Deacetylation degree (A) is calculated by the following equation.

[0014]

However, a is the weight of the sample (g), and f is 0.
The titer of 1N-caustic soda aqueous solution, b is the titer (ml) of 0.1N-caustic soda aqueous solution.

The bone morphogenetic protein of the present invention is a substance which acts on undifferentiated mesenchymal cells extracellularly and induces its genetic trait to chondrocytes or osteoblasts (chondrocyte induction, osteoinduction). BMP (Born Morphogenetic Pro), which can be obtained by, for example, a method of separating and purifying from Dunn osteosarcoma.
tein: Takaoka, K., Biomedical Research, 2 (5) 466-471 (19
81)) is known. Separately, BDGF (Born Der
ived Growth Factor: Canalis, E., Science, 210,1021 (198
0)), CDF (Cartilage Derived Factor: Anderson, H.
C., Am.J.Pathol, 44,507 (1964)), SGF (Skeletal G
rowth Factor: Farly, JR, Biochemistry, 21,3508 (198
2)), OGF (Osteogenic Factor: Amitani, K., Calcif.
Tiss. Res., 17, 139 (1975)) and the like are known.

Kunio Takaoka et al., Orthopedic and Disaster Surgery, 26
(10), 1451 (1983) also discloses a method for extracting and purifying the same, and any of these osteogenic factors can be obtained by a known method. In addition, human bones, bovine bones, or bone morphogenetic factors obtained by genetic recombination can also be used.

The chitin used in the present invention can be used as a powder as it is, or can be used as a molded product. The molded body can be prepared by dissolving chitin powder in a solvent to form a dope and solidifying the same, and examples thereof include fibers, fibrils, films, porous bodies, and microbeads.

The solvent is preferably a mixture of halogenated hydrocarbon and trichloroacetic acid, N-methylpyrrolidone or dimethylacetamide and lithium chloride. As the coagulant, water, alcohols, ketones and the like are preferable.

Chitin can be molded with these solvents and coagulants by existing methods. For example, when the chitin fiber is wet-molded, chitin is dissolved in the above solvent and filtered through a mesh stainless net to obtain a transparent dope.
Further, the dope is fed under pressure, discharged from a nozzle into hot water to be solidified, and wound on a bobbin.

In order to produce a molded product of deacetylated chitin, a chitin molded product having a low degree of deacetylation is first prepared,
Deacetylation is performed by treating the molded body with concentrated alkali, and the degree of deacetylation can be adjusted depending on the conditions. The treatment method may be the same as the method for deacetylating chitin, for example, by immersing the molded body in a concentrated alkaline solution and allowing it to stand at room temperature or high temperature for a certain period of time. The degree of deacetylation can be freely selected by changing the temperature and the standing time.

In the case of chitosan, which is chitin having a high degree of deacetylation, it can be easily molded by a method of removing the acid after dissolving it in an acid solution and molding. Here, an acetic acid solution is preferable as the acid solution. As a molded product of chitosan, for example, in the case of molding a chitin film, chitosan can be dissolved in a 2% acetic acid aqueous solution and cast on a glass plate to evaporate water and acetic acid at room temperature or high temperature to produce a film. .

Next, the method for producing the chitin composition containing an osteogenic factor may be any method as long as the osteogenic factor can be adsorbed, contained or chemically bound to chitin. For example, as a simple method, a method of suspending a bone morphogenetic protein in a solvent, immersing chitin in this suspension, and then drying it can be mentioned. The solvent used at this time may be one that does not inactivate the osteogenic factor. For example, there is a method of suspending an osteogenic factor in water, dipping and stirring chitin in the suspension, removing water and drying. Alternatively, a method of kneading an osteogenic factor when molding a chitin molded product may be used.
For example, a method of injection-molding a chitin dope with a bone-forming factor simultaneously, or a method of simultaneously containing a chitin-dope with a bone-forming factor and removing the solvent by a method such as drying. Using a cross-linking agent such as aldehyde,
There is also a method of binding chitin or a chitin-molded body to an osteogenic factor. There is also a method of chemically or physically binding an osteogenic factor to chitin or a chitin molded body. Furthermore, there is a method of injecting a bone morphogenetic protein into a chitin molded body with a syringe or the like.

The chitin molded product containing the bone morphogenetic protein obtained by the above-mentioned production method can be used as it is, or can be powdered again and used. It is also possible to use a reaction accelerator such as a catalyst for the reaction of.

The concentration of the osteogenic factor in the chitin composition of the present invention can take various values depending on the field and application,
The concentration is not limited to a certain range and can be prepared at any concentration, but it is desirable that the weight percentage of the bone morphogenetic protein is 0.01% or more based on the total weight. For example, when the weight of the bone morphogenetic protein with respect to the total weight is expressed as a percentage (percentage), it is 25 when a large amount is required.
% Or more, and if a small amount is required, the content may be 5% or less.

[0026]

EXAMPLES The present invention will be described in more detail below with reference to examples.

Example 1 Crude chitin powder (manufactured by Shin Nippon Kagaku Co., Ltd.) was crushed to 100 mesh, treated with 1N hydrochloric acid at 4 ° C. for 1 hour, and further heated in a 3% caustic soda aqueous solution at 90 ° C. for 3 hours. The calcium and protein contained in the crude chitin powder were removed by treatment. Deacetylation degree of this chitin powder is 5.2%
Met. This was dissolved in a mixed solution of dimethylacetamide and lithium chloride at 6 wt% and filtered through a 1480 mesh stainless net to prepare a transparent dope. This dope was put in a tank, defoamed under reduced pressure, and then pumped under pressure with a gear pump, spitted into hot water at 80 ° C from a nozzle with a diameter of 0.07 mmφ and 500 holes, and then solidified at 10 m / mi.
It was wound on a bobbin at a speed of n to produce a chitin fiber having a single yarn denier of about 1 d (total denier = about 500 d). When this chitin fiber was treated in a 20 wt% caustic soda aqueous solution at 95 ° C. for 2 hours, the degree of deacetylation was 41% and the fiber was highly water resistant. On the other hand, bone morphogenetic protein (BMP) 25
A solution was prepared by suspending mg in 5 ml of water, and the above fiber was immersed in the suspension and freeze-dried to obtain a BMP-containing chitin fiber. The BMP content at this time was 5.5 wt%. This fiber was wrapped around a ceramic having a diameter of 2 mm and a total length of 15 mm, transplanted into the interdorso muscle of a mouse, and the implant was taken out after 3 weeks. As a result, bone tissue was formed around the implant.

Example 2 The same chitin powder as in Example 1 was dissolved in a mixed solution of dimethylacetamide and lithium chloride in an amount of 2 wt% to prepare a transparent dope. Polyvinyl alcohol was added to this dope, and after sufficiently stirring, it was molded into a cylindrical shape having a diameter of 3 mm and a total length of 15 mm, and depolyvinyl alcohol was removed in hot water to prepare sponge chitin. On the other hand, bone morphogenetic protein (BMP) 1
A liquid in which 5 mg was suspended in 3 ml of water was prepared, and the above-mentioned spongy chitin was dipped and freeze-dried to obtain a BMP-containing chitin molded body. The BMP content rate at this time was 7.1%. This molded body was transplanted into the intermuscular region of the back of a mouse, and the graft was taken out after 5 weeks. As a result, the chitin molded body was replaced with bone tissue, and its wet weight was 18.9 mg.

Example 3 The spongy chitin of Example 2 was used. Bone morphogenetic protein (BMP) was injected into this by a syringe to obtain a BMP-containing chitin molded body. The BMP content rate at this time was 13.4.
%Met. About 50 mm ^{3 of} this compact was transplanted into the muscles of the back of the mouse, and the graft was taken out after 5 weeks.
It was mg.

[0030]

INDUSTRIAL APPLICABILITY The chitin composition of the present invention has excellent affinity with living bodies, no foreign body reaction in living bodies, excellent sustained release of bone morphogenetic factors, and exhibits sufficient bone forming ability.

Claims (1)

[Claims] 1. A chitin composition comprising chitin containing an osteogenic factor.