JPH0797715A - Production of collagen fiber - Google Patents

Abstract

PURPOSE:To obtain collagen fiber having an extremely low content of residual organic solvent free from foreign matters. CONSTITUTION:In producing collagen fiber by regenerating solubilized collagen, collagen fiber formed into a fibrous state is cleaned with an organic solvent which is soluble in water but does not dissolve the collagen fiber or a mixed solution of the organic solvent and water and solvent is removed at <=80 deg.C at 30-98% relative humidity for >=10 minutes to give the objective collagen fiber.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for producing collagen fibers which can be used as a hemostatic material.

[0002]

2. Description of the Related Art As a hemostatic method during surgery, a compression method,
There are applications of physiologically active substances such as ligation method, electrocoagulation method, thrombin and fibrin glue method. Ligation or electrocoagulation is generally used for arterial bleeding with a clear bleeding point, and for venous bleeding, compression alone is sufficient and hemostasis is easy. However, these hemostatic methods may not be effective for bleeding from solid organs and capillary bleeding, and hemostasis is particularly difficult when bleeding tends to occur in liver failure or cardiovascular surgery. . In such a case, a locally absorbed hemostatic material that accelerates the blood coagulation reaction just by contacting the bleeding surface, forms a thrombus promptly and prevents bleeding, not only shortens the operation time but also causes re-bleeding after the operation. It is also effective because it also contributes to safe postoperative management.

For this purpose, various local hemostatic materials made of oxidized cellulose have been developed and clinically applied.
Although it has the advantage of being inexpensive and excellent in operability,
Since it is not a material derived from a living body, it has a drawback that it is slowly absorbed in a living body and causes a hemolytic reaction. A local hemostatic agent using collagen, which is a protein capable of minimizing the reaction, has been actively used clinically because it has its own physiologically active action and also has a high hemostatic effect.

[0004] Currently available collagen local hemostatic materials include those in which collagen fine fibers are formed into flakes and collagen sponges are formed into flat plates, but the former is in the form of flakes. Since it is washed away with blood and scattered, it cannot be expected to have a hemostatic effect, and it is easy to be charged with static electricity, and when it is used, it is easy to adhere to hands and tweezers, which is a difficulty in operation. On the other hand, with regard to the latter, since it is a flat plate, it does not have sufficient adhesiveness to the wound surface of a complicated shape, and pressure hemostasis cannot be performed.
Similar to the former, the hemostatic effect cannot be expected so much.

As a solution to the above-mentioned drawbacks, there is a fibrous local hemostatic material comprising atelocollagen regenerated from solubilized collagen. The hemostatic material has a sufficient hemostatic ability and, at the time of solubilization, is antigenic. Since the determinant telopeptide part has been removed, it is a hemostatic material that is safe in vivo after use, is rapidly decomposed and absorbed, and has good operability.

Such a collagen fiber is manufactured as follows. For example, insoluble collagen obtained from fresh cowhide is enzymatically treated with a protease, pepsin, to digest telopeptide, and a solution of solubilized atelocollagen is produced. A spinning stock solution having a collagen concentration of 6 wt% can be dissolved, and atherocollagen fibers can be formed into a fibrous shape by a wet spinning method in which a spinning solution of 20 wt% ammonium sulfate is spun.

However, the collagen fiber thus produced contains residual salts of various salts provided in the production, or foreign substances mixed in from the equipment and materials used in the process. Although these are extremely small amounts, they cause a problem in the use of providing collagen fibers, for example, as a hemostatic material. Therefore, it is important to effectively remove these residues and contaminants by some method.
As a method, it is effective to wash with an organic solvent that dissolves in water but does not dissolve collagen fibers. However, after this cleaning step, the problem that the used organic solvent remains 5 wt% or more becomes a problem, and it is necessary to effectively remove the organic solvent.

It was thought that such removal of the organic solvent could be easily achieved by heat treatment in the vicinity of the boiling point of the organic solvent, but in reality, it not only causes alteration and morphological change of the collagen fiber, but also unexpectedly. Also, 1 to 10 wt% of organic solvent
Also has the problem of remaining. Moreover, the residue of the organic solvent does not change even when heated or depressurized, and is independent of the pressure. Also, to avoid the deterioration of collagen fibers,
Similar to the heating method, the so-called freeze-drying method of cooling to near the crystallization temperature of the organic solvent under reduced pressure is almost the same as the removal of the organic solvent.

[0009]

That is, one of the causes of the above-mentioned phenomenon is considered to be that the dense layer is formed on the surface layer of the collagen fiber, which hinders the diffusion of the organic solvent. Therefore, the present inventors need to use a mechanism other than simple diffusion in combination, and as a result of the study based on the idea of promoting diffusion of the organic solvent from the inside of the fiber by using molecular water as a carrier of the organic solvent, In the removal of organic solvent, the humidity which was not considered in the past was controlled, and collagen fiber was placed under the condition of a constant relative humidity range. The organic solvent was drastically reduced to 0.05 wt% or less, and The present invention was completed by discovering an effect that rapidly decreased and far exceeded expectations. Moreover, according to the present invention, since the temperature is not a major controlling factor, it has been found that it has a practically extremely large feature that it does not cause the deterioration and deformation of collagen fibers by heating. The present invention is based on such knowledge, and an object thereof is to obtain collagen fibers free from foreign matter and having an extremely small residual organic solvent.

[0010]

According to the present invention, when solubilized collagen is regenerated to produce collagen fibers, the collagen fibers formed into a fibrous shape are dissolved in water but do not dissolve the collagen fibers. After washing with a solvent or a mixed liquid of an organic solvent and water, a solvent removal treatment is carried out under the conditions of a temperature of 80 ° C. or lower and a relative humidity of 30 to 98% for a time of 10 minutes or more. On the way.

In the present invention, the fibrous shaped collagen fibers are obtained by a known spinning method by spinning a spinning dope containing solubilized collagen as an aqueous solution into a coagulating solution of a concentrated aqueous solution of salts. In the present invention, collagen fibers spun by a known method and shaped into fibers are washed with an organic solvent or a mixed solution of an organic solvent and water.

As the organic solvent, one having a solubility in water has a large penetration into the collagen fiber tissue and has a great effect of cleaning impurities. Further, the use as a mixed solution with water further enhances the cleaning effect. However, it goes without saying that the organic solvent should not dissolve the collagen fibers. From these viewpoints, as the organic solvent preferably used in the present invention, for example, alcohols,
Ketones, glycols, cellosolves, tetrahydrofuran, dimethyl sulfoxide, dimethylformamide, dimethylacetamide, dioxane, pyrrolidone,
Acetic acid etc. can be mentioned.

Among them, alcohols are suitable solvents when collagen fibers are used as medical products such as hemostatic materials. Examples of such alcohols include methanol, ethanol, isopropanol, n-
Propanol, n-butanol, isobutanol, se
c-Butanol, tert-butanol or mixtures thereof may be mentioned as effective solvents. The ketones methyl ethyl ketone and tetrahydrofuran are also effective solvents.

In the organic solvent removal treatment of the present invention, the temperature must be 80 ° C. or lower. At a temperature above 80 ° C, deformation, dissolution, partial adhesion and melting of collagen fibers are caused. The low temperature has no problem, but the forced cooling is disadvantageous in terms of energy cost.

The relative humidity is an important constituent factor in the organic solvent removal treatment of the present invention, and in the present invention, the relative humidity is required to be 30 to 98%. Relative humidity is 3
If it is less than 0%, the effect of removing the solvent is poor, and if the humidity exceeds 98%, deformation, dissolution, partial adhesion and melting of the fiber are caused, which is not preferable. The time may be selected as appropriate until reaching the target residual concentration level, but is set to 10 minutes or longer, preferably 30 minutes or longer.

In the present invention, prior to the solvent removal treatment step, collagen fibers are subjected to a crosslinking treatment such as a crosslinking agent treatment, a heat treatment and an energy irradiation treatment to crosslink at least a part of the fibers to control the solubility of the fibers. Doing so does not hinder the effect of the present invention.

[0017]

EXAMPLES The present invention will be specifically described below with reference to examples. In the examples, as a method for analyzing the residual organic solvent in the collagen fiber, a method in which collagen fiber is dispersed or dissolved in water, and distillation and a distillate are quantified by a gas chromatograph method is adopted.

Example 1 Insoluble collagen obtained from fresh cowhide was enzymatically treated with proteolytic enzyme pepsin, and solubilized atelocollagen was adjusted to pH with an aqueous hydrochloric acid solution.
The collagen fibers prepared by the wet spinning method using ammonium sulfate as a coagulating liquid are washed with a mixed liquid of isopropanol and water in a weight ratio of isopropanol / water = 80/20, desalted, and then centrifugally dehydrated. Was used to remove excess isopropanol, which was dried at room temperature in vacuo at room temperature.
The isopropanol concentration in this fiber was 10 wt%. This collagen fiber was subjected to solvent removal treatment for 3 hours in a forced circulation atmosphere of air having a temperature of 30 to 80 ° C. and a relative humidity of 70 to 90%. The obtained collagen fiber had a residual isopropanol concentration in the fiber as shown in Table 1, and the isopropanol was effectively removed.

[0019]

[Table 1]

Example 2 Collagen fibers obtained by washing, dealcoholization by centrifugation and vacuum drying in Example 1 were crosslinked by heat treatment at 100 ° C. and 1 Torr or less for 5 hours. The residual isopropanol concentration in this crosslinked collagen fiber was 7 wt%. This fiber was subjected to a solvent removal treatment for 10 minutes to 3 hours in an atmosphere in which air having a temperature of 30 ° C. and a relative humidity of 90% was forcedly circulated. The obtained collagen fiber had the residual isopropanol concentration in the fiber as shown in Table 2, and the isopropanol was effectively removed.

[0021]

[Table 2]

Example 3 Except that ethanol was used instead of isopropanol in Example 1, washing was performed in the same manner as in Example 1, excess ethanol was removed by centrifugation, and vacuum drying was performed at room temperature. did. The ethanol concentration in this fiber was 4 wt%. This collagen fiber was subjected to dealcoholization treatment for 3 hours in an atmosphere in which air having a temperature of 30 to 80 ° C. and a relative humidity of 70 to 90% was forcedly circulated.
The obtained collagen fiber had a residual ethanol concentration in the fiber as shown in Table 3, and ethanol was effectively removed.

[0023]

[Table 3]

Example 4 The collagen fiber obtained by the crosslinking treatment in Example 2 was treated with Na ₂ CO ₃ · 10H ₂ at room temperature.
An aqueous solution in equilibrium with the solid phase of O was placed in a container having a volume 20 times the volume of this aqueous solution, and the humidity was adjusted to 24, 48 in a natural convection atmosphere of air whose relative humidity was adjusted to about 90% relative humidity.
The solvent was removed for 72 hours. The collagen fiber obtained is
The residual isopropanol concentration in the fiber was as shown in Table 4, and the isopropanol was effectively removed.

[0025]

[Table 4]

(Example 5) In Example 1, washing was performed in the same manner as in Example 1 except that ethanol was used instead of isopropanol, excess ethanol was removed by centrifugation, and vacuum drying was performed at room temperature. did. The ethanol concentration in this fiber was 4 wt%. The collagen fiber was subjected to a solvent removal treatment for 30 minutes in this atmosphere by forcedly blowing air with a temperature of 30 ° C. and a relative humidity of 80 to 95% from one direction. The obtained collagen fiber had a residual ethanol concentration of 0.05 wt% in the fiber, and it was found that ethanol was effectively removed.

(Example 6) After removing excess tetrahydrofuran by centrifugation in the same manner as in Example 1 except that tetrahydrofuran was used instead of the mixed solution of isopropanol and water. , Vacuum dried at room temperature. The tetrahydrofuran concentration in this fiber was 3 wt%. This collagen fiber at a temperature of 30
3 in an atmosphere with forced circulation of air at ℃ and 90% relative humidity
The solvent was removed for a period of time. The obtained collagen fiber had a residual tetrahydrofuran concentration of 0.05 wt% in the fiber, and the tetrahydrofuran was effectively removed.

Comparative Example 1 Collagen fibers obtained by dealcoholating by washing and centrifugation in Example 1 were vacuum dried at room temperature and 1 Torr or less for 5 hours. The obtained collagen fiber had a residual isopropanol concentration of 11 wt% in the fiber, and isopropanol was not removed by vacuum drying.

Comparative Example 2 Collagen fibers obtained by dealcoholating by washing and centrifugation in Example 1 were vacuum dried at a temperature of −45 ° C. and 1 Torr or less for 5 hours. The collagen fiber obtained had a residual isopropanol concentration in the fiber of 10 wt%.

Comparative Example 3 The collagen fiber obtained by washing and centrifuging to dealcoholize in Example 1 was dried with hot air at a temperature of 100 ° C. for 2 hours. The collagen fiber obtained has a residual isopropanol concentration of 7 wt% in the fiber.
Met.

Comparative Example 4 Collagen fibers obtained by dealcoholating by washing and centrifugation in Example 1 were vacuum dried at a temperature of 100 ° C. and 1 Torr or less for 4 hours. The obtained collagen fiber had a residual isopropanol concentration in the fiber of 7 wt%.

(Comparative Example 5) Collagen fibers obtained by dealcoholation by washing and centrifugation in Example 3 were vacuum dried at a temperature of 100 ° C and 1 Torr or less for 4 hours. The obtained collagen fiber had a residual ethanol concentration in the fiber of 4 wt%.

Comparative Example 6 Collagen fibers obtained by washing and centrifuging to dealcoholize in Example 1 were dried in hot air at a temperature of 100 ° C. for 2 hours and treated at a temperature of 90 ° C. and a relative humidity of 98% for 3 hours. . Although this treated product had a reduced residual isopropanol concentration, it melted and did not assume a fiber morphology.

[0034]

INDUSTRIAL APPLICABILITY The collagen fiber according to the present invention is a safe collagen fiber having a very small amount of residual organic solvent, and is preferably used as a collagen hemostatic material and can be a fiber aggregate of any shape. Since the handling problems of the material are improved and the adhesion to the hemostatic surface is improved, it is possible to provide a hemostatic material that can be used quickly for hemostasis and has a high hemostatic effect. When at least a part of the fibers are cross-linked, a hemostatic material that is safe and has higher hemostatic ability can be provided.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Takashi Takigawa 4-chome Sunadabashi, 1-chome, Higashi-ku, Nagoya, Aichi Prefecture 60-72, Product Development Laboratory, Mitsubishi Rayon Co., Ltd. (72) Kaoru Terasawa 4-chome, Sunadabashi, Higashi-ku, Aichi Prefecture No. 60 Mitsubishi Rayon Co., Ltd. Product Development Laboratory

Claims (4)

[Claims] 1. When regenerating solubilized collagen to produce collagen fibers, a fibrous shaped collagen fiber is dissolved in water but does not dissolve the collagen fiber, or an organic solvent or an organic solvent and water. A method for producing collagen fibers, which comprises washing with a mixed solution, and then subjecting to a solvent removal treatment under an atmosphere of a temperature of 80 ° C. or less and a relative humidity of 30 to 98% for a time of 10 minutes or more. 2. The enzyme-treated or alkali-treated atelocollagen is used as the solubilized collagen.
A method for producing the collagen fiber described. 3. An organic solvent comprising at least one selected from the group consisting of methanol, ethanol, isopropanol, n-propanol, n-butanol, tert-butanol, sec-butanol, acetone, methyl ethyl ketone and tetrahydrofuran. Item 2. A method for producing a collagen fiber according to item 2. 4. The method for producing collagen fibers according to claim 1, 2 or 3, wherein the collagen fibers are crosslinked prior to the solvent removal treatment.