JPH06197946A - Locally absorbing hemostatic material - Google Patents

Abstract

PURPOSE:To obtain a local hemostatic material made of collagen which has a sufficient blood stopping function while allowing safe and quick absorption in vivo after use and moreover, provides high operability by accomplishing a cross linking of at least a part of a collagen molecule composing a fiber by heat. CONSTITUTION:An insoluble collagen obtained from a fresh bovine skin is treated by pepsin to obtain an atherocollagen with telopeptide made soluble while being digested. This solution is dissolved into a hydrochloric acid aqueous solution to obtain an atherocollagen fiber by a wet spinning method ammonium sulfate as a coagulation liquid. The fiber is desalted and dewatered by methanol and cut to a length of 50mm after being dried to be separated by an air blowing. Then, a thermal treatment is performed at a specified temperature to obtain a cotton0like product of the collagen fiber. The length of the fiber is 3-70mm, the diameter thereof 10-70mum and a specific capacity 2-80cm<2>.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a locally absorbable hemostatic material composed of a cotton-like material of collagen fibers used in the surgical field, and particularly to bleeding from a solid organ or capillary bleeding. The present invention relates to a locally absorbable hemostatic material that can be quickly and effectively applied.

[0002]

2. Description of the Related Art As a hemostatic method during surgery, a compression method,
There are ligation method, electrocoagulation method, and application of physiologically active substances such as thrombin and fibrin glue. Ligation or electrocoagulation is generally used for arterial bleeding with a clear bleeding point, and if venous bleeding, compression is sufficient and hemostasis is easy. However, these hemostatic methods may be ineffective for bleeding from a solid organ or capillary bleeding, and hemostasis is particularly difficult when bleeding tends to occur in liver failure or cardiovascular surgery. In such a case, a locally absorbable hemostatic material that accelerates the blood coagulation reaction just by contacting the bleeding surface, quickly forms a thrombus, and prevents bleeding, not only shortens the operation time but also causes re-bleeding after surgery. It is also effective because it also contributes to safe postoperative management. For this purpose, various topical hemostatic agents using oxidized cellulose as a material have been developed and clinically applied.However, although they have the advantage of being inexpensive and excellent in operability, they are not materials of biological origin, so in vivo It has a drawback that it is slow to be absorbed and induces a strong hemolytic reaction and a foreign body reaction, and in recent years, since the allergic reaction and the foreign body reaction can be minimized because the antigenicity is low and it is safely absorbed by the body, The local hemostatic agent using collagen is its own physiologically active action (adhesion and aggregation of platelets, action of platelet factor III released from the aggregated platelets to accelerate the blood coagulation system and form thrombus). It has been widely used clinically due to the fact that it has (and therefore has a high hemostatic effect).

A collagen-made topical that is currently in practical use
As a hemostatic material, fine particles of collagen were made into flakes.
There is a flat plate type of thing or collagen sponge
However, for the former, it is an antigenic determinant of collagen.
Shows antigenicity in vivo due to residual telopeptides
It must be removed after use and is flaky.
Therefore, it has a hemostatic effect because it is washed away by blood and scatters.
You can not expect it, and it is easy to be charged with static electricity,
There is a difficulty in operation that it easily adheres to the tweezers.
On the other hand, for the latter, remove the telopeptide
Some are made of atherocollagen, but are flat.
Therefore, the adhesion to the wound surface with a complicated shape is not sufficient.
In addition, it is not possible to stop pressure bleeding.
Can't expect much.

As a solution to the above-mentioned drawbacks, a cotton-like local hemostatic material comprising atelocollagen has been announced. For example, pig-skin-derived atelocollagen is spun and dried into a cotton shape (Shimizu et al., Artificial organ 19 (3), 1
235 (1990)) or spun collagen treated with a cross-linking agent, washed and freeze-dried to give a cotton-like shape and fine cracks on the surface (Japanese Patent Laid-Open No. 4-61862). However, the former uses soluble atelocollagen as it is, and therefore, when used for hemostasis, it may absorb blood and significantly reduce the strength of the fiber to block blood flow. It is difficult to expect a sufficient hemostatic effect. On the other hand, in the latter, the hemostatic action of collagen is impaired because the crosslinking treatment with a crosslinking agent is performed prior to freeze-drying, and in order to completely remove the unreacted crosslinking agent from the viewpoint of safety. There was a problem that a cleaning operation was required.

[0005]

DISCLOSURE OF THE INVENTION The present invention provides a hemostatic material that solves the above-mentioned problems of the conventional collagen hemostatic material, that is, it has a sufficient hemostatic ability and is decomposed and absorbed safely in vivo after use. It is an object of the present invention to provide a collagen local hemostatic material which has excellent operability.

[0006]

As a result of repeated studies for such purpose, the present invention has been achieved. That is, the present invention is
A cotton-like material composed of atelocollagen fibers regenerated from solubilized collagen, the diameter of the fibers being 10 to 70 μm.
m, the length of the fiber is 3 to 70 mm, and at least a part of the collagen molecule constituting the fiber is crosslinked by heat.

In the present invention, the solubilized collagen is a solubilization treatment with a proteolytic enzyme or an alkali solubilization treatment, and at the same time as the solubilization, an operation for removing telopeptide which is an antigenic determinant of collagen is performed. The applied color
It is Gen. The origin of the collagen used as the raw material of the present invention is not particularly limited, but generally, the skin, bone, cartilage, tendon, organ of mammals (for example, human, bovine, porcine, rabbit, sheep, rat etc.) Collagen obtained from, for example, is used. Also, a collagen-like protein obtained from birds, fish and the like can be used.

In general, collagen which has been solubilized with a proteolytic enzyme has a higher activity on platelets than that which has been solubilized with an alkali, and therefore collagen which has been solubilized with a proteolytic enzyme is used. -Preference is given to using gen.

Hemostasis by a collagen hemostatic material is performed by physically suppressing the blood that springs out first and then exerting the hemostatic action of collagen itself. Therefore, in order to exert a sufficient hemostatic effect, a hemostatic material should be used. The strength of the constituent fibers is an important factor. Regarding the strength of the fiber, the thicker the diameter of the constituent fiber is, the larger the diameter of the constituent fiber becomes.
˜70 μm is preferable, and 15 to 45 μm is preferable.

Regarding the length of the fiber, it can be obtained by regenerating soluble collagen so that the fiber can be of any length, but if it is too long, it can be dispersed sufficiently when it is made into a cotton shape. If the length is too short, the fibers will not easily get entangled with each other, and the fibers will be scattered when the hemostatic material is applied, and the strength against blood pressure cannot be obtained.
m is good, and preferably 5 to 50 mm.

The locally absorbable hemostatic material of the present invention swells when it contains blood, and the whole volume increases. As a result, a compression and hemostatic effect is exerted on the tissue surrounding the application site. The compressive hemostatic effect is exhibited only when the hemostatic material has some strength.
The fiber obtained from the solubilized collagen has a significant degree of swelling, and the strength decreases with the swelling. Therefore, in the present invention, at least a part of the collagen molecules constituting the fiber is crosslinked by heat. , While maintaining the hemostatic action originally possessed by collagen, moderate swelling properties as a hemostatic material (the fiber has a dense surface structure, but immediately absorbs blood to cause volume expansion, We decided to give it strength (the effect of completely blocking the blood flow and achieving hemostasis early), which brings about a pressure hemostatic effect on the bleeding site.

Conventionally, a method using a cross-linking agent such as aldehydes and diepoxides has been used in the case of cross-linking collagen. However, in the case of collagen treated with the cross-linking agent, the activity of collagen on platelets is high. Therefore, improvement of hemostatic ability cannot be expected (the heat treatment does not easily inactivate the reactivity of collagen with platelets). In addition, the method using a cross-linking agent cannot stop the reaction by the degree of cross-linking that leaves an appropriate degree of swelling as a hemostatic agent, and since the cross-linking agent used is often toxic, unreacted There are many problems such as a cleaning operation for removal.

The thermal crosslinking treatment is preferably carried out in the range of 50 to 200 ° C. If the temperature is lower than 50 ° C, the crosslinking will be insufficient or the crosslinking will not be performed. On the other hand, if the temperature is higher than 200 ° C, the collagen is remarkably denatured and the swelling property and the physiological activity are impaired. Considering both manufacturing and quality factors, the temperature is more preferably 60 to 180 ° C, and particularly preferably 95 to 150 ° C.

As another physical property that should be possessed as the locally absorbable hemostatic material, the specific volume of the cotton-like material of collagen fiber is also important in terms of handleability of the hemostatic material and adhesion to the wound site. The specific volume of the fiber is easy to shape when applied (the hemostatic effect can be maximized by changing the shape of the hemostatic material according to the wound site with various shapes and evenly adhering it. It is better to be able to do so), but there is a limit to the fact that the fibers can be properly entangled with each other, and if it is too high, it will be difficult to completely stop hemostasis because the fibers will scatter and gaps will be created during application. Therefore, 20 to 80 cm ³ / g is preferable, and 4 is preferable.
It is about 0 to 70 cm ³ / g.

Here, the specific volume is determined as follows (hereinafter, the cotton-like material of collagen fiber is described as a reference, but the flaky material is also measured in the same manner). The obtained cotton-like material of collagen fibers was put in a standard state (20
Approximately 1 g is taken under (° C, 65% RH) and accurately weighed (measured weight: Wg). Under standard conditions, it is uniformly filled in a transparent plastic cylinder with an inner diameter of 35 mm. Next, a flat disk with a diameter of 30 mm and a weight of 5.0 g
It is placed on the cotton-like material prepared in step 1, and a weight of 50 g is placed on the cotton-like material for 30 seconds, and then the weight is removed and left for 30 seconds. This operation is repeated 3 times, and then the height of the filled cotton-like material is measured at 3 locations in the circumferential direction (the average value is Hmm.
And). The specific volume was calculated for three samples according to the following formula,
The average value is adopted. Specific volume (cm ³ / g) = ((35/20) ² × π × H /
10) / W

[0016]

【Example】

Example 1 Insoluble collagen obtained from fresh cowhide was treated with a proteolytic enzyme (pepsin; the same applies below) to digest telopeptide and obtain a solubilized atelocollagen solution. Next, this was dissolved in a hydrochloric acid aqueous solution adjusted to pH 2 (collagen concentration: 6%), and an atelocollagen fiber was obtained by a wet spinning method using 20% ammonium sulfate as a coagulating liquid. The obtained atelocollagen fiber is desalted with methanol, dehydrated, dried and cut into a length of 50 mm, and the fiber is dispersed by air blow (10 minutes at a wind speed of 30 m / s), and then 105 ° C. Heat treatment (3 hours) was performed to obtain a cotton-like material of collagen fibers. The physical properties of the obtained cotton-like material are as follows: fiber length: 5 to 50 mm, fiber diameter: 1
It was 5 to 45 μm and the specific volume was 60 cm ³ / g. still,
The shape of the fiber surface of the obtained cotton-like material was as shown in FIG. 1 and FIG. 2 (observation result by scanning electron microscope: JSM-840A type manufactured by JEOL Ltd.).

Example 2 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the heat treatment temperature was 60 ° C. (24 hours). The physical properties of the obtained cotton-like material were the same as in Example 1.

Example 3 Example 1 was repeated except that the heat treatment was performed at 180 ° C. (10 minutes).
A cotton-like material of collagen fibers was obtained in the same manner as in. The physical properties of the obtained cotton-like material were the same as in Example 1.

Example 4 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the fiber was not dispersed by air blowing. The physical properties of the obtained cotton-like material are as follows: fiber length: 30-50
mm, fiber diameter: 15 to 45 μm, specific volume: 28 cm ³
/ G.

Example 5 The conditions for fiber dispersion treatment by air blowing were 40 m / s.
A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the wind speed was changed to 60 minutes. The physical properties of the obtained cotton-like material are as follows: fiber length: 5 to 50 mm, fiber diameter: 15 to 45
μm, specific volume: 78 cm ³ / g.

Example 6 Collagen was prepared in the same manner as in Example 1 except that alkali solubilization treatment (using a mixed solution of sodium sulfate, sodium hydroxide and monomethylamine) was carried out in place of the enzyme treatment of insoluble collagen. A fibrous material was obtained. The physical properties of the obtained cotton-like material were the same as in Example 1.

Example 7 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that fresh pig skin was used instead of fresh cowhide. The physical properties of the obtained cotton-like material were the same as in Example 1.

Comparative Example 1 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that heat treatment was not performed. The physical properties of the obtained cotton-like material were the same as in Example 1.

Comparative Example 2 Except that an alkali solubilization treatment (using a mixed solution of sodium sulfate, sodium hydroxide and monomethylamine) was carried out in place of the enzyme treatment of insoluble collagen, and no heat treatment was carried out. A cotton-like material of collagen fibers was obtained in the same manner as in Example 1. The physical properties of the obtained cotton-like material were the same as in Example 1.

Comparative Example 3 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the treatment was performed with glutaric aldehyde instead of the heat treatment. The physical properties of the obtained cotton-like material were the same as in Example 1.

Comparative Example 4 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the spinning conditions were changed. The physical properties of the obtained cotton-like material are
The fiber length was 5 to 50 mm, the fiber diameter was 7 to 9 μm, and the specific volume was 65 cm ³ / g.

Comparative Example 5 A cotton-like material of collagen fibers was obtained in the same manner as in Example 1 except that the spinning conditions were changed. The physical properties of the obtained cotton-like material are
Fiber length: 5 to 50 mm, fiber diameter: 80 to 100 μm,
Specific volume: 35 cm ³ / g.

Comparative Example 6 A flaky collagen fiber material was obtained in the same manner as in Example 1 except that the length after cutting was set to 3 mm. The physical properties of the obtained flaky product are as follows: fiber length: 0.5 to 1 m
m, fiber diameter: 15 to 45 μm, specific volume: 18 cm ³ /
It was g.

Comparative Example 7 Insoluble collagen obtained from fresh cowhide was treated with a proteolytic enzyme to digest telopeptide and a solubilized atelocollagen solution was obtained. Next, this was dissolved in a hydrochloric acid aqueous solution adjusted to pH 2 (collagen concentration: 3%), and collagen fibers were obtained by a wet spinning method using saturated sodium sulfate as a coagulating liquid. Next, this was treated with a glutaraldehyde solution and then washed with water to remove inorganic salts and unreacted glutaraldehyde. Next, the fiber was cut into a length of 50 mm and freeze-dried to obtain a cotton fiber of collagen fiber. The physical properties of the obtained cotton-like material are as follows: fiber length: 4
0 to 50 mm, fiber diameter: 10 to 30 μm, specific volume: 1
It was 7 cm ³ / g.

[0030]

[Effect of the invention] Heparin is added at 100 u / kg before the start of the experiment.
A systemically-administered mixed-breed dog is opened under general anesthesia, and only the spleen capsule is exactly peeled off with a scalpel to a size of 1 cm x 1 cm. Next, 0.1 g of each of the collagen cotton-like material and the flaky material obtained in each of the above Examples and Comparative Examples was applied to the release surface and pressed for 30 seconds, and then the blood loss was measured at 1-minute intervals. (The gauze dried for 1 minute was applied to the bleeding surface, and the radius of the circle drawn by the blood sucked by the gauze at that time was defined as the bleeding amount. Hereinafter, this index is referred to as Bleeding D
egree: BD. The initial value of BD is BD _{t = 0} ,
BD is BD _{t = t,} and the value obtained according to the following equation is the hemostasis rate: HR. In addition, hemostasis was completed when the blood did not soak into the gase). HR (%) = (1− (BD _{t = t} / BD _{t = 0} )) × 100 The data obtained when 10 cases were applied are shown in Table 1. In the table, 0.1 g of the collagen cotton-like material and the flaky material obtained in each of the above Examples and Comparative Examples was processed according to the shape of the release surface, and applied to the release surface. The results of measurement of the time required for (reprocessing time) are also shown.

[0031]

[Table 1]

From the results, it is clear that the hemostatic collagen cotton-like material according to the present invention is excellent in both hemostatic ability and handleability (in this experiment, hemostasis was completed at the initial stage (up to 2 minutes) of hemostasis). In the non-experimental example, the exuding blood partly pushed the hemostatic material away (because the fibers constituting the hemostatic material cannot maintain the strength enough to withstand the blood pressure), and thus hemostasis became difficult. It is extremely important that hemostasis is completed by 2 minutes).

The cotton-like material of collagen fiber according to the present invention is
It is a fiber aggregate having an appropriate length and specific volume obtained by regenerating solubilized collagen, and at least a part of collagen molecules constituting the fiber is thermally crosslinked,
The effect of suppressing hemorrhage, which is the first step of hemostasis, is improved, and sufficient hemostatic ability is exhibited without impairing the original hemostatic action of collagen. Moreover, since it has an appropriate specific volume, it can be easily processed according to the shape of the bleeding site, can be used quickly for hemostasis, and has good adhesion to the bleeding surface and a high hemostasis effect.

[Brief description of drawings]

FIG. 1 is a micrograph showing the shape of the surface of the atelocollagen fiber constituting the locally absorbable hemostatic material of the present invention (magnification: 1
000 times). FIG. 2 is a micrograph (magnification: 7,000 times) of the surface further enlarged.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Toru Natsume Toru Natsume 3-3 Midorigahara, Tsukuba-shi, Ibaraki Central Research Institute, Nippon Ham Co., Ltd. (72) Toshikazu Makihara 3-3 Midorigahara, Tsukuba, Ibaraki Japan Ham Central Research Institute, Inc. (72) Inventor Masanori Akasaka 4-chome, Sunadabashi, Higashi-ku, Aichi Prefecture Mitsubishi Rayon Co., Ltd. Product Development Laboratory (72) Inventor, Keiki Sakakibara 2-3-19 Kyobashi, Chuo-ku, Tokyo Within Mitsubishi Rayon Co., Ltd.

Claims (3)

[Claims] 1. A cotton-like material comprising atelocollagen fibers regenerated from solubilized collagen, wherein the collagen fibers have a diameter of 10 to 70 μm and a fiber length of 3 to 70 mm.
A locally absorbable hemostatic material characterized in that at least a part of collagen molecules constituting the fiber is crosslinked by heat. 2. The locally absorbable hemostatic material according to claim 1, wherein crosslinking by heat is carried out at 50 to 200 ° C. 3. The specific volume of the cotton-like material is 20 to 80 cm ³ /
The locally absorbable hemostatic material according to claim 1 or 2, which is g.