JP2007161521A - Fine carbon fiber dispersion - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a fine carbon fiber dispersion excellent in dispersion stability and useful especially for being introduced into a living body. <P>SOLUTION: The carbon fiber dispersion is made by dispersing carbon fiber of 0.001-30 mass% in polyethylene glycol. By adding fine carbon fiber directly into polyethylene glycol and shaking, the carbon fiber is very stably and well dispersed. When the dispersion is diluted with various media, especially with aqueous media such as water, the state where the fine carbon fiber is stably and finely dispersed in these media is kept for a long time. Therefore, the carbon fiber dispersion is suitable especially for being introduced into a living body. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a fine carbon fiber dispersion. More specifically, the present invention relates to a fine carbon fiber dispersion that is excellent in dispersion stability and particularly useful for introduction into a living body.

  In recent years, fine carbon fibers such as carbon nanostructures typified by carbon nanotubes (hereinafter also referred to as “CNT”) have attracted attention.

  The graphite layer constituting the carbon nanostructure is a substance having a regular, six-membered ring arrangement structure, and chemically, mechanically and thermally stable properties as well as its unique electrical properties. Such fine carbon fibers are expected to be applied not only to various industrial uses but also to pharmacological and medical uses.

  However, on the other hand, such fine carbon fibers are already agglomerated at the time of production, and it is difficult to stably and uniformly disperse, for example, even if dispersed in a solvent such as physiological saline, It was difficult to introduce fine carbon fibers into the living body as fine particles.

  In addition, Patent Document 1 shows a composition in which various fibers are dispersed in a polyol as a cosmetic application. However, the composition is intended for external application, and there are a wide variety of fibers. It contains fiber types and is not intended for its dispersion characteristics.

Recently, the problem of health damage such as emphysema caused by asbestos has been widely taken up, and as the use of fine carbon fiber expands, it is necessary to demonstrate such a safety aspect. Yes. For this reason, for example, an experiment animal such as a mouse is placed in an atmosphere in which fine carbon fibers are suspended for a predetermined time, and attempts are made to put fine carbon fibers into the lungs of the experimental animal by exhalation. Even if fine carbon fibers cannot be introduced into the glass, and even if they can be introduced, it is difficult to make the introduction amount constant, and such safety evaluation is extremely difficult. there were.
JP 2000-344627 A

  Accordingly, an object of the present invention is to provide a fine carbon fiber dispersion which is excellent in dispersion stability and is particularly useful for introduction into a living body.

  Another object of the present invention is to provide an established experimental method as a method for evaluating the safety of fine carbon fibers.

  This invention which solves the said subject is a carbon fiber dispersion characterized by being made to disperse | distribute in a polyol in the ratio of 0.001-30 mass% of the whole.

  The present invention also shows the carbon fiber dispersion, wherein the polyol is at least one selected from the group consisting of polyethylene glycol and propylene glycol.

  The present invention further shows the carbon fiber dispersion, wherein the carbon fiber is further diluted with an aqueous medium so that the carbon fiber is in a ratio of 0.001 to 30% by mass.

  Furthermore, the present invention is the carbon fiber dispersion which is used for introduction into a living body.

  The present invention that solves the above-mentioned problems also introduces the carbon fiber dispersion into a tissue or organ of an experimental animal, so that the carbon fiber is quantitatively placed in the body of the experimental animal. This is a carbon fiber safety evaluation method characterized in that follow-up is observed.

  The present invention also provides a method for incising the neck of a laboratory animal by a surgical procedure to secure an airway, and injecting the carbon fiber dispersion described above into the airway, thereby introducing the carbon fiber into the lung of the laboratory animal. 1 shows a carbon fiber safety evaluation method for suturing the surgical site and then observing the state of the experimental animal.

  In the present invention, the skin of an experimental animal is incised by a surgical procedure, and the carbon fiber dispersion is injected subcutaneously, thereby implanting carbon fiber in the subcutaneous tissue of the experimental animal, and then suturing the surgical site. Thereafter, a carbon fiber safety evaluation method for observing the state of the experimental animal is shown.

  The present invention further relates to a method for evaluating the safety of carbon fiber in which the experimental animal is a mouse.

  In order to solve the above-mentioned problems, the present inventors have intensively studied, and by adding fine carbon fiber directly to a polyol such as polyethylene glycol and stirring or shaking, it is easy, very stable and good. Even if the carbon fiber is dispersed in the medium and then diluted with an aqueous medium such as various media, particularly water, the fine carbon fiber can maintain a stable and finely dispersed state in these media for a long time. The present invention has been found and reached the present invention. In addition, this shows a clearly different behavior in terms of fine dispersion and dispersion stability compared to the case where fine carbon fibers are dispersed in a system in which a polyol component is previously added to an aqueous medium. In addition, as is well known in the art, a good dispersion system can be obtained without using a dispersant or dispersion stabilizer such as a surfactant.

  Furthermore, in the carbon fiber safety evaluation method of the present invention, since the carbon fiber dispersion having excellent dispersibility as described above is used, the carbon fiber is quantitatively and safely introduced into the tissues or organs of experimental animals. can do. In particular, when a small animal such as a mouse is used, even if it is injected directly into the airway of the experimental animal and carbon fiber is introduced into the lung, the experimental animal will die of suffocation immediately after injection because the carbon fiber is clogged in the airway Since it is possible to survive the experimental animal for a long time after the operation, it is possible to introduce a predetermined amount of carbon fiber into the lung of the experimental animal and observe the subsequent course Become. For this reason, it is an extremely excellent method for evaluating the safety of carbon fibers.

  Hereinafter, the present invention will be described in detail based on preferred embodiments.

<Carbon fiber dispersion>
The carbon fiber dispersion according to the present invention is characterized in that carbon fibers are dispersed in a polyol at a ratio of 0.001 to 30% by mass of the whole.

A) Dispersion medium In the present invention, the polyol as the dispersion medium includes various organic compounds containing at least two free hydroxy groups as known in the art. Specifically, for example, propylene glycol, butylene glycol, Isoprene glycol, pentylene glycol, hexylene glycol, glycerol, panthenol, polyethylene glycol, for example, polyethylene glycol having an average molecular weight of about 180 to 10,000, polypropylene glycol, for example, polypropylene glycol having an average molecular weight of about 60 to 200, etc. be able to.

  Of these, polyethylene glycol, particularly polyethylene glycol having an average molecular weight of about 190 to 1500 and propylene glycol are desirable. Polyethylene glycol and propylene glycol are preferable dispersion media from the viewpoint of biosafety.

  Further, the viscosity of the dispersion medium is not particularly limited, but it is desirable that the dispersion medium be in a liquid state at 25 ° C. ± 5 ° C.

B) Carbon Fiber Next, the carbon fiber used in the present invention is not particularly limited, and various conventionally known carbon fibers can be used. A structure having a structure, in which at least one of the three-dimensional dimensions of the structure is in a nanometer region, for example, a fine carbon fiber of about 0.4 to 150 nm, preferably 15 to 100 nm. is there.

  As this carbon six-membered ring arrangement structure, a sheet-like graphite graphene sheet can be exemplified, and further, for example, a structure in which a five-membered ring or a seven-membered ring is combined with a carbon six-membered ring, etc. Can be included.

  More specifically, as the fine carbon fiber, for example, a single-walled carbon nanotube having a diameter of about several nanometers formed by rounding a graphene sheet into a cylindrical shape, or a multilayer in which cylindrical graphene sheets are laminated in a direction perpendicular to the axis Examples include carbon nanotubes (multi-walled carbon nanotubes), carbon nanohorns in which the ends of single-walled carbon nanotubes are closed in a conical shape, and carbon nanohorn aggregates in which the carbon nanohorn is a spherical aggregate having a diameter of about 100 nm. it can. Further, carbon onions having a carbon six-membered ring arrangement structure, fullerenes and nanocapsules in which a five-membered ring is introduced into the carbon six-membered ring arrangement structure, and the like are included. In the present invention, these fine carbon fibers can be used alone or in a mixture of two or more.

  As a method for producing such a fine carbon fiber, a catalytic metal and a gaseous or liquid carbon-containing compound are put into a high-temperature (600 ° C. to 1300 ° C.) reaction furnace, and they are mutually exchanged in the reaction furnace region. A CVD (Chemical Vapor Deposition) method is possible in which the raw material is pyrolyzed or catalytically decomposed to carbon by contact and grown.

  Also, the CVD method differs from the catalyst introduction method, in which metal fine particles are previously supported on a carrier and introduced into a reaction furnace as a solid, and a gaseous or liquid carbon-containing compound is introduced into the reaction furnace. In addition, there is a CCVD (Catalytic Chemical Vapor Deposition) method in which fine carbon fibers are obtained by bringing them into contact with metal fine particles previously introduced and supported on a carrier.

  In general, a catalytic metal is used in the CVD method. In an as-grown fine carbon fiber, these metals or metal ions may be contained in the fine carbon fiber. When the carbon fiber dispersion according to the present invention is used for, for example, introduction into a living body or preparation of a product that may come into contact with a living body or tissue, there is a possibility of toxicity due to such a metal or metal ion. Therefore, it is desirable to use carbon fibers obtained by removing these metal components as much as possible after heat treatment, for example, heat treatment at 1500 to 3000 ° C., more preferably 2000 to 3000 ° C., after the production of fine carbon fibers. .

  In addition, as a production method for obtaining fine carbon fibers, a voltage is applied between two electrodes after lightly contacting two graphite electrodes in an atmosphere of low-pressure argon gas or hydrogen gas, and separated by several mm. The arc discharge method that sublimates the graphite electrode to produce fine carbon fibers by causing arc discharge to occur, and the graphite mixed with the metal catalyst is irradiated with visible pulsed laser light, and the graphite is evaporated to produce fine particles. Various production methods such as a laser evaporation method for obtaining carbon fibers can be mentioned, and any production method can be used as long as the above-described fine carbon fibers can be obtained.

  In addition, in such applications for introduction into a living body, the fine carbon fibers to be used should be appropriately sterilized using, for example, steam sterilization, chemical sterilization using ethylene oxide or formaldehyde, ultraviolet sterilization, radiation sterilization, etc. It is desirable to sterilize in advance.

C) Method for Preparing Carbon Fiber Dispersion The carbon fiber dispersion of the present invention can be prepared by blending a predetermined amount of carbon fiber as described above in a polyol as a dispersion medium and stirring or shaking the mixture. Is possible.

  The amount of the carbon fiber arranged in such a polyol is about 0.001 to 30% by mass, more preferably about 0.001 to 10% by mass of the entire composition. When the blending amount is in the range of 0.001 to 30% by mass, a stable dispersion containing a sufficient amount of carbon fibers in a finely dispersed state is formed with good fluidity. is there.

  The method of stirring or shaking treatment is not particularly limited, and various known methods, specifically, automatic or manual shakers, magnetic stirrers, ultrasonic vibrators, other paddles, etc. It can be performed by using a stirrer equipped with various stirrers such as blades, a grinder, etc. Among them, in particular, the dispersion treatment using a magnetic stirrer destroys the structure of the carbon fiber itself. And desirable dispersion state can be formed.

  The carbon fiber dispersion according to the present invention can be prepared as described above, and the dispersion in which the carbon fibers are dispersed in the polyol in this way can be prepared by using various media, In particular, it is possible to dilute in an aqueous medium. Even when the carbon fiber is diluted to a ratio of 0.001 to 30% by mass with such an aqueous medium, the carbon fiber is finely dispersed in the obtained dilution system. Can keep stable.

  When the carbon fiber dispersion according to the present invention is used for in vivo introduction, examples of the aqueous medium used for dilution include water such as distilled water and deionized water, physiological saline, phosphate buffer, serum for culture, and the like. It is desirable that the medium be highly biosafe.

  In addition, when diluting in the aqueous medium as described above, it is possible to further blend various conventionally known dispersants, dispersion stabilizers and the like with the carbon fiber dispersion according to the present invention. Even if a dispersant, a dispersion stabilizer, etc. are not added, sufficient dispersion stability can be exhibited, and adverse effects due to surfactants used as a dispersant, for example, effects from the viewpoint of biocompatibility, etc. In view of the above, there are many embodiments in which it is desirable not to add these dispersants, dispersion stabilizers and the like.

<Uses of carbon fiber dispersion>
As described above, the carbon fiber dispersion according to the present invention can be preferably used for various uses, particularly for in vivo introduction, because the carbon fibers are uniformly finely dispersed and exhibit a stable dispersion state.

Although it is not particularly limited as a biointroduction application, in addition to the carbon fiber safety evaluation method as described later, for example, as carbon fiber, pharmacologically active substances such as various drugs, antibodies, fluorescent substances, etc. As a carrier of a drug delivery system using a carbon fiber bonded or supported with a carbon fiber, which is introduced into a living body to locally give these pharmacologically active substances;
Utilizing the good thermal conductivity of carbon fiber, as a heating medium for thermal treatment that generates heat by infrared irradiation or the like after being introduced into a predetermined site in the living body;
Utilizing the good dispersibility of fine carbon fibers in the carbon fiber dispersion, the fine carbon fibers are introduced into the capillaries or peripheral blood vessels in the vicinity of local sites such as pathological sites in the living body, and inside the capillaries or peripheral blood vessels. As a plugging material for emergency hemostasis using a catheter, as well as a material used for occlusion for occluding a blood vessel with fine carbon fiber and removing necrosis of a local site such as a pathological site ahead of it;
As a radiation or chemical marker for detecting carbon fiber with a suitable labeling substance such as a radioactive substance or a fluorescent substance and introducing it into a predetermined site in the living body;
Contrast agent for magnetic resonance used for diagnosis and treatment by nuclear magnetic resonance (NMR) imaging or biological magnetic resonance imaging (MRI) apparatus by enclosing a proton-bearing substance such as water in a liquid state in the hollow portion of carbon fiber As;
As a scaffold material for cell culture in regenerative medicine using the good dispersibility of fine carbon fibers in carbon fiber dispersions;
Etc. can be illustrated.

  In addition, various medical devices utilizing the good biocompatibility of carbon fiber, in particular, various injection materials such as injection needles, indwelling needles, indwelling catheters, artificial organs, artificial bones, etc. that can come into contact with tissues or cells in vivo It can also be suitably used for preparing a coating agent used for the purpose of modifying the surface of a medical device.

<Safety evaluation method for carbon fiber>
Since the carbon fiber dispersion according to the present invention exhibits very good and stable dispersibility in the dispersion medium composed of polyol as described above, the carbon fiber dispersion is in a very fine cavity. However, it can pass stably without causing blockage.

  For this reason, if the carbon fiber dispersion according to the present invention is used, carbon fiber can be introduced not only into in vitro experiments but also directly into tissues such as subcutaneous tissues of experimental animals or organs such as lungs. It is possible to establish an effective in vivo method for evaluating the safety of carbon fibers.

  That is, in the carbon fiber safety evaluation method according to the present invention, first, the carbon fiber to be tested is prepared in the form of a carbon fiber dispersion using a polyol as described above, By injecting the carbon fiber dispersion into the tissues or organs, the carbon fibers are placed in these tissues or organs, and then the survival state of the experimental animals, pathological changes such as tissues, blood, etc. are followed up It is.

  By using the carbon fiber dispersion according to the present invention, since the burden on the experimental animal at the time of carbon fiber introduction is greatly reduced, the mortality rate of the experimental animal due to the introduction procedure itself is extremely low, Since quantitative introduction of carbon fibers is possible, reliable and stable evaluation is possible.

  As an experimental animal to be used for the test, it can be applied to any kind of animal that is usually used. Although not particularly limited, for example, mice, rats, rabbits, dogs, goats, sheep, pigs, monkeys and the like can be used. Among these, it is particularly desirable to use a mouse from the viewpoints of existence of abundant known data, ease of subculture, availability, gene operability, and the like.

  Although it does not specifically limit as a mouse | mouth, For example, an immunodeficient mouse | mouth, especially ASW, A / He, AKR, BALB / c, B10LP, C17, C3H, C57BL, C57, CBA, DBA, DDD, I Preferred examples include nude mice such as / st, NC, NFR, NFS, NFS / N, NZB, NZC, NZW, P, RIII, and SJL.

  In addition, as a more specific introduction method to a tissue or organ, for example, as shown below, introduction into a lung of a laboratory animal, implantation into a subcutaneous tissue, and the like can be preferably exemplified.

  That is, the introduction into the lung is achieved by incising the neck of the experimental animal by a surgical procedure to secure the airway, and injecting the carbon fiber dispersion according to the present invention described above into the airway. This can be done by suturing the surgical site after introducing the carbon fiber inside.

  In the introduction into the lung, when the carbon fiber dispersion according to the present invention was used, even in an animal species having a very thin airway such as a mouse, it was included in the introduced carbon fiber dispersion. Since the carbon fiber has extremely good dispersibility, it is very unlikely that an accident in which the carbon fiber is clogged in the airway or the like and the mouse is suffocated immediately after the procedure will occur. Therefore, it is possible to prepare a subject in which a predetermined amount of carbon fiber is introduced into the lung with a good survival rate, and it is possible to perform animal experiments over a long period of time related to the safety of carbon fiber.

  In the safety evaluation method according to the present invention, first, the carbon fiber to be inspected is blended in a polyol such as polyethylene glycol as described above at a ratio of 0.001 to 30% by mass of the carbon. A fiber dispersion is prepared.

  Next, the carbon fiber dispersion is diluted with water, physiological saline, or the like as necessary so that the carbon fibers are in a proportion of 0.001 to 30% by mass.

  Then, an experimental animal, for example, a mouse subjected to the test is anesthetized in advance as necessary, and then the neck is opened by a surgical technique to secure an airway.

  Subsequently, an injection needle, for example, in the case of a mouse, an injection needle having a thickness of about 27G is punctured into this airway, and a predetermined amount of carbon fiber dispersion is introduced from a syringe or the like connected to the injection needle. In order to reduce the possibility of suffocation of experimental animals by holding the injection needle connected to the syringe in the airway for a relatively long time, an indwelling needle with an open end is previously placed in the airway. It is desirable to reduce the burden on the laboratory animal by allowing the laboratory animal to breathe through the opening of the indwelling needle and not allowing exhalation to enter the lung for only a short time after the carbon fiber dispersion injection. .

  Finally, when the injection of the predetermined amount of carbon fiber dispersion is completed, the incision site is immediately sutured to obtain a subject into which the predetermined amount of carbon fiber has been introduced into the lung. After the operation, for example, after 1 to 2 hours of follow-up, and if survival is confirmed, the subject is subjected to a subsequent carbon fiber safety test.

  The safety test can be performed by, for example, tissue observation by subject anatomy at predetermined time intervals thereafter, analysis of collected blood, survival rate after a predetermined time, immune reaction, pathological examination, genetic examination, protein expression examination, etc. it can.

  In this safety evaluation method, the control (control) is not particularly limited. For example, a polyol such as polyethylene glycol used as a dispersion medium without adding carbon fiber, or water, physiological An experimental animal in which a diluted body with saline or the like is introduced into the lung by the same procedure as described above can be used.

  In the implantation into the subcutaneous tissue, an appropriate part of the experimental animal, for example, the skin of the part such as the back is surgically incised, and a predetermined amount of the above-mentioned carbon fiber dispersion is injected subcutaneously into the incised part. Thereafter, the incision site can be sutured.

  Since the carbon fiber dispersion to be used, follow-up after the operation, safety test, and the like are the same as those in the case of the introduction into the lung described above, description thereof will be omitted.

  EXAMPLES Hereinafter, although an Example demonstrates this invention in more detail, this invention is not limited to the following Example at all.

  In the following, each physical property value was measured as follows.

(1) X-ray diffraction Using a powder X-ray diffractometer (JDX3532, manufactured by JEOL Ltd.), carbon fibers after the annealing treatment were examined. The Kα ray generated at 40 kV and 30 mA in a Cu tube is used, and the surface spacing is measured according to the Gakushin method (the latest carbon material experiment technology (analysis and analysis), edited by the Carbon Materials Society of Japan). Used as internal standard.

(2) Raman spectroscopic analysis The Raman spectroscopic analysis was performed using an instrument (LabRam 800) manufactured by Horiba Joban Yvon Co., Ltd. and using an argon laser at 514 nm.

(Synthesis Example 1)
Fine carbon fibers were synthesized using toluene as a raw material by the CVD method.

  The synthesizer is shown in FIG.

  A mixture of ferrocene and thiophene was used as a catalyst, and the reaction was performed in a hydrogen gas reducing atmosphere. Toluene and the catalyst were heated to 375 ° C. together with hydrogen gas, supplied to the production furnace, and reacted at 1200 ° C. with a residence time of 8 seconds. The atmospheric gas was separated by a separator and recycled. The hydrocarbon concentration in the feed gas was 9% by volume.

  The tar content of the fine carbon fiber of the synthesized intermediate substance (first intermediate substance) was 10%.

  Next, the fiber was heated to 1200 ° C. and held for 30 minutes to perform hydrocarbon separation treatment, and further subjected to high-temperature heat treatment at 2500 ° C. An apparatus for hydrocarbon separation and high temperature heat treatment process is shown in FIG.

  Moreover, FIG. 1 demonstrated above is an electron micrograph of the fine carbon fiber which carried out the hydrocarbon separation process at 1200 degreeC. From this figure, it can be seen that the graphene sheets constituting the fine carbon fibers are not continuous and have a patch shape.

  An electron micrograph of the obtained fine carbon fiber after high-temperature heat treatment at 2500 ° C. is shown in FIG.

From this figure, fine carbon fibers having a unique structure can be confirmed. As a result of SEM observation, the diameter of the produced fiber varied to some extent, and was 10 to 60 nmφ, and the specific surface area was 29 m ² / g. The magnetoresistance value has a negative value with respect to the magnetic flux density and has a negative slope with respect to the change in the magnetic flux density (the first derivative is negative with respect to the magnetic flux density B). The measured I _D / I _G was 0.05.

  Moreover, the obtained fine carbon fiber was measured using the X-ray-diffraction apparatus. For comparison, graphite was also measured using an X-ray diffractometer. The X-ray diffraction chart obtained from the measurement results is shown in FIG. 3, but the fine carbon fiber obtained in Synthesis Example 1 has a weak peak intensity, and is compared by making it 10 times.

  From this result, the plane spacing of the obtained fine carbon fiber was 3.388 angstroms.

Example 1
The carbon fiber synthesized in Synthesis Example 1 is blended in 10 ml of polyethylene glycol (average molecular weight 190 to 210) so as to be 1% by mass of the whole, and is stirred for 3 minutes at a rotational speed of 1000 rpm using a magnetic stirrer. The carbon fiber dispersion was obtained by processing. FIGS. 6 (a) and 6 (b) show an appearance photograph and a micrograph (magnification 40 times) showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion.

Example 2
The carbon fiber dispersion prepared in Example 1 was diluted with distilled water to obtain a carbon fiber dispersion containing 5% by mass, 10% by mass, and 20% by mass of the carbon fibers. FIG. 7 shows a photograph showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion.

Comparative Example 1
In advance, distilled water was added to polyethylene glycol (average molecular weight 190 to 210) at the same rate as in Example 2, and the resulting mixture was mixed at the same rate as in Example 2 to Synthesis Example 1. The carbon fiber synthesized in the above is blended and stirred using a magnetic stirrer under the same conditions as in Example 1, and the carbon fiber structure is contained in 5% by mass, 10% by mass, and 20% by mass for comparison. A carbon fiber dispersion was obtained. A photograph showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion is shown in FIG.

Example 3
The carbon fiber structure synthesized in Synthesis Example 1 was blended with 10 ml of propylene glycol so as to be 1% by mass of the whole, and stirred with a magnetic stirrer at a rotation speed of 1000 rpm for 3 minutes. A fiber dispersion was obtained. FIGS. 9A and 9B show an outer appearance photograph and a micrograph (magnification 40 times) showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion.

Example 4
The carbon fiber dispersion prepared in Example 2 was diluted with distilled water to obtain a carbon fiber dispersion containing 5% by mass, 10% by mass, and 20% by mass of the carbon fiber structure. A photograph showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion is shown in FIG.

Comparative Example 2
In advance, distilled water was added to propylene glycol at the same rate as in Example 4, and the carbon fiber synthesized in Synthesis Example 1 at the same rate as in Example 4 to the resulting mixture. And a carbon fiber dispersion for comparison containing 5% by mass, 10% by mass, and 20% by mass of the total carbon fiber structure by stirring with a magnetic stirrer under the same conditions as in Example 1. Obtained. A photograph showing the dispersion state observed after 5 minutes of standing of the obtained carbon fiber dispersion is shown in FIG.

  As is apparent from the results shown in FIGS. 6 to 11, in the dispersions of Examples 1 to 4 according to the present invention, the carbon fiber structures are stably finely dispersed in the medium without agglomeration. In contrast to those observed in Comparative Examples 1 and 2, the carbon fiber structure was agglomerated with almost no dispersion in the aqueous medium, although it was almost identical in composition to Examples 2 and 4. It was in a state of floating near the interface.

Examples 5 to 6, Comparative Example 3 and Reference Example 1
75 5-week-old female BALB / c mice were purchased from SLC and raised in a sterile animal facility. The mice were allowed to stabilize for 1 week in the facility and the experiment was started when the age was 6 weeks old. The mice were randomly divided into four groups (for Examples 5, 6, Comparative Example 3 and Reference Example 1 (Comparative Control)).

  After anesthesia with Frocene just before the procedure, the neck was incised, the airway was secured, a 27G injection needle was punctured, and prepared in Example 1 or Example 2 from a syringe connected to the injection needle 0.05 ml of carbon fiber dispersion was injected. Immediately after the injection of the predetermined amount of carbon fiber dispersion was completed, the incision site was sutured to obtain a subject mouse in which the predetermined amount of carbon fiber was introduced into the lung.

  The survival rate of mice 30 days after the operation was 96% in Example 3 in which the dispersion of Example 1 was introduced, and 96% in Example 4 in which the dispersion of Example 2 was introduced. Survival rate of 96% in mice of Reference Example 1 (comparative control group) in which an unblended polyethylene glycol aqueous solution (polyethylene glycol concentration is the same as in Example 2) was injected into the respiratory tract by the same procedure, the significant difference was There wasn't.

  Furthermore, the subject mice in which a predetermined amount of carbon fiber was introduced into the lungs were bred for a long period of time (2 months or more), but the survival rate of the mice was very high, both 96% and 96%. Met.

  After the test period, the mouse was sacrificed and sacrificed. As a result, no abnormalities such as emphysema were observed in the lung tissue, and the bio-integrity of the carbon fiber obtained in Synthesis Example 1 was confirmed.

  On the other hand, in Comparative Example 3, carbon fibers were introduced into the lungs in the same manner as in Examples 3 to 4, except that the carbon fiber dispersion to be injected was prepared in Comparative Example 2. Tried.

  As a result, immediately after the dispersion injection, all the mice died in a state considered to be due to suffocation, and it was impossible to obtain a subject mouse in which a predetermined amount of carbon fiber was introduced into the lung.

Example 7
Experimental method (preparation of carbon fiber dispersion)
Four types of carbon fibers (SWNT, MWNT-I, MWNT-II, cap laminated CNT) having the characteristics shown in Table 1 below were prepared.

  Each preparation is as follows.

<SWNTs>
Single-walled carbon nanotubes are produced by catalytic CVD and optimized purification methods (Endo M, Hayashi T, Kim YA, Muramatu H, Ezaka M, Watts PCP, et al. The possible route to large-scale production of SWNTs through J Nanos Nanotec 2004; 4 (1/2): 132-135.).

<MWNT-I and MWNT-II>
Two types of multi-walled carbon nanotubes with different diameters were selected from samples obtained by catalytic CVD with controlled reaction conditions (reaction temperature, amount of organometallic compound, flow rate).

(Endo M, Kim YA, Hayashi T, Nishimura K, Matusita T, Miyashita K, et al. Vapor-grown carbon fibers (VGCFs): Basic properties and their battery application. Carbon 2001; 39: 1287-1297;
Endo M, Kim YA, Hong SH, Matushita T, Takeda T, Hayashi T, et al. Structural characterization of carbon nanofibers obtained by hydrocarbon pyrolysis. Carbon 2001; 39: 2003-2010; and
Kim YA, Hayashi T, Endo M, Kaburagi Y, Tsukada T, Shan J, et al. Synthesis and structural characterization of thin multi-walled carbon nanotubes with a partially facetted cross section by a floating reactant method.Carbon 2005; In press. checking ... )
The as grown MWNT-II was heat-treated in a graphite furnace at 2800 ° C. under argon to remove the polycyclic aromatic hydrocarbons and iron compounds.

<Cap laminated CNT>
Floating reaction method: obtained in a continuous process using catalytic metal, hydrogen sulfide as cocatalyst and natural gas as carbon feedstock (Endo M, Kim YA, Fukai T, Hayashi T, Oshida K, Terrones M, et al. Structural Characterization of cup-stacked type nanofibers with an entire hollow core. See Appl Phys Lett 2002; 80 (7): 1267-1269.).

  The resulting tube exhibits a unique morphology (conical frusto-stacked morphology and high chemical reactivity on most of the outer and inner surfaces of the open edge.

^(a) The diameter and length of the multi-walled carbon nanotube were measured by FE-SEM, while the diameter and length of the single-walled carbon nanotube were measured by RBM (radial breathing mode) of Raman spectrum.
^(b) d ₍₀₀₂₎ is the interlayer distance from the XRD pattern. .
^(c) Specific surface area was measured by N ₂ absorption.
^{(d) The} true density was measured with a pycnometer.
^{(e) The} R value is a value obtained by dividing the D band intensity in the Raman spectrum by the G band intensity.
^{(f) The} iron content was measured by atomic absorption method.
^{(g) The} heat treatment was performed in argon.

  Prior to the experiment, all carbon fiber samples were sterilized with formaldehyde gas for 24 hours, evaporated at room temperature for 24 hours, and embedded in the subcutaneous (cell) tissue under the skin on the back.

  Next, each carbon fiber sample was blended in 10 ml of (average molecular weight 190 to 210) so as to be 1% by mass of the whole, and stirred with a magnetic stirrer at 1000 rpm for 3 minutes. A carbon fiber dispersion was obtained.

(Experimental animals)
180 female BALB / c mice (age 5 weeks) were purchased from SLC and bred in a sterile animal facility. The mice were allowed to stabilize for 1 week in the facility and the experiment started when the age was 6 weeks.

  The mice were randomly divided into five groups (30 each) for comparison, SWNT, MWNT-I, MWNT-II, and cap-stacked CNT. Furthermore, it was divided into 6 subgroups on the day of sampling.

Sampling day: After implantation, 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months Sampling was performed by collecting blood from the heart and (cell) tissue.

(Implantation surgery)
Under Fluxen anesthesia, the skin on the back of each mouse was incised about 1 cm, and 2 mg of each carbon fiber dispersion was implanted subcutaneously and sutured. As a control, 2 ml of physiological saline was used. On the designated sampling day, blood and tissue were sampled under deep fluosane anesthesia. Note that the sampled mouse is sacrificed.

  Blood was collected by puncturing the heart for measurement by flow cytometry, and skin tissue including the muscle layer was collected for pathological examination.

(Flow cytometry)
Sampled blood was stained with CD4 ⁺ and CD8 ⁺ antibodies (BD Bioscience Pharmingen, USA) to sort T lymphocytes on a flow cytometer (FACS Clibur, Becton Dickinson, USA).

  Stained immune cells were analyzed by CeIIQuest software (Becton Dickinson).

(Histology and immunity)
According to traditional histology, immediately after sacrifice, (cell) tissue was treated with 70% ethanol, incorporated in paraffin, cut into 5 μm sections, and stained with hematoxin and eosin.

In addition, using one of the serial sections, endogenous peroxidase was blocked with 3% H ₂ O ₂ for 10 minutes, and anti-mouse CD45R antibody (BD Bioscience Pharmingen, USA) and anti-mouse Mac-3 antibody ( BD Bioscience Pharmingen, USA) for 60 minutes at room temperature. Slides were washed and applied with a biotinylated polyclonal rat-IgG1 / 2a antibody (BD Bioscience Pharmingen, USA). After washing three more times with phosphate buffer, the sections were incubated with peroxidase-conjugated streptavidin (BD Bioscience Pharmingen, USA) for 60 minutes, stained with diambenzidine, and counterstained with hematoxin.

(Statistical analysis)
Effective differences were calculated using two-tailed student's t-test (P <0.05.). Data showed ± SE.

Experimental results (weight)
All mice survived throughout the study. There was no change in body weight compared to the control (3 months).

  Therefore, it was shown that the carbon fiber safety test method according to the present invention works effectively.

(Blood analysis)
Comparative control The comparative control showed that CD4 ⁺ T cell values were 30.7 ± 4.9%, 36.3 ± 2.2%, 27.2 ± 3.3%, 32.4 ± 0.6 at 1, 2, 3 weeks, 1 month, 2 months and 3 months, respectively, after implantation. %, 40.9 ± 2.31% and 28.9 ± 3.9%, and CD8 ⁺ T cell values are 11.1 ± 0.9%, 11.6 ± 0.4%, 16.3 ± 2.9%, 10.1 ± 0.14%, 12.3 ± 1.4% and 7.4 ± 0.6 %Met.

SWNT
FIG. 12 shows the time course of peripheral T cells after SWNT subcutaneous implantation. CD4 ⁺ T cells embedded with SWNT were significantly higher at 26.3 weeks after implantation (46.3 ± 2.2%) and at 3 months (44.2 ± 3.9%) than the control (non-implanted) (FIG. 12 (a)). In addition, SWNTs CD8 ⁺ T cells showed a significant decrease compared to controls at 1 week (7.5 ± 0.1%) and 3 weeks (11.0 ± 0.7%) after implantation (FIG. 12 (b)). Therefore, it should be noted that SWNTs-embedded T cells showed relatively higher CD4 ⁺ / CD8 ⁺ values than controls at 1 and 3 weeks after implantation (3.6 ± 0.3 and 2.8 ± 0.3 respectively). (FIG. 12C).

  To confirm the changes in peripheral T cells, a histopathological study of samples from 1 week to 1 month after SWNTs subcutaneous implantation was performed (FIG. 13). In one week, a loose (loose) space representing the presence of exudative fluid (edema) was clearly observed between SWNTs and tissues.

  This space became smaller two weeks after implantation. However, cell infiltration by SWNT occurred during this period. And 3 weeks after implantation, granulation seed tissue due to cell infiltration was observed. The granulation seed encapsulating this SWNT mass became hard one month after implantation. Therefore, there was no evidence of cell infiltration into the SWNT mass. Perhaps this is due to SWNT's own dense packing structure.

MWNT-I
CD4 ⁺ T cell values were significantly reduced at 1 week (17.8 ± 4.9%) after implantation and greatly increased at 2 weeks (48.6 ± 2.2%) (FIG. 14 (a)). Compared to the control, a significant decrease in CD8 ⁺ T cells was also observed at 1 week (6.9 ± 0.5%) and 3 weeks (8.6 ± 0.7%) after implantation (FIG. 14 (b)). Therefore, relatively high CD4 ⁺ / CD8 ⁺ values were observed only 2 weeks after implantation (3.8 ± 0.1) (FIG. 14 (c)). As can be seen in FIG. 15, MWNT caused cell invasion one week after implantation without edema-like appearance. The granulated tissue became dense and hard over time between the nanotube masses.

  Finally, after 3 weeks after implantation, some particles were phagocytoseed.

MWNT-II
Relatively low values of CD4 ⁺ T cells were observed at 1 week (16.4 ± 0.8%) and 3 weeks (13.7 ± 1.9%) after implantation. In the period of 2 weeks (53.5 ± 0.8%) after implantation, a high value of CD4 ⁺ was detected (FIG. 16 (a)). Similarly, a significant decrease in CD8 ⁺ T cells was observed at 1 week (8.2 ± 0.8%) and 3 weeks (7.8 ± 0.9%) after implantation. Moreover, it should be noted that a significant increase in CD8 ⁺ T cells occurred even 3 months after implantation (FIG. 16 (b)). Therefore, the value of CD4 ⁺ / CD8 ⁺ was much lower (2.1 ± 0.2) than the control. However, in this sample, a significantly high value of CD4 ⁺ / CD8 ⁺ was observed 2 weeks after implantation (4.7 ± 0.3) (FIG. 16 (c)). From the histopathological study of the tissue in which MWNT-II was implanted, not only slight cell infiltration but also slight granulation seed formation was observed in the 1 week after implantation as well as a decrease in exudation. Cell infiltration was in close proximity to the embedded mass 2 weeks after implantation. However, cell infiltration did not occur in the central space of the mass even over one month. On the other hand, the mass was tightly closed with granulation seed tissue as shown in FIG.

Cap laminated CNT
The changes in CD4 ⁺ T cells in this group were 1 week (18.2 ± 1.4%) higher after implantation compared to controls, and at 2 weeks (53.5 ± 0.8%) and 3 months (40.1 ± 3.2%), It was almost doubled (FIG. 18 (a)). On the other hand, compared to the control, a sudden decrease in CD8 ⁺ T cells was observed at 1 week (6.6 ± 0.6%) and 3 weeks (6.6 ± 0.9%) after implantation (FIG. 18 (b)). Therefore, high values of CD4 ⁺ / CD8 ⁺ were obtained at 2 weeks (4.2 ± 0.1), 3 weeks (3.3 ± 0.3) and 3 months (4.8 ± 0.5) after implantation (FIG. 18 (c)). As shown in FIG. 19, this tube caused slight cell infiltration with an edema-like appearance as early as one week after implantation. As time passed, the granulated tissue became smaller and harder between the tube masses. It is noteworthy that highly dispersible tubes (spider webs) produce lower cell infiltration than massive tubes.

(Comparison of immunohistochemical studies using Mac-3 antibody and CD45R antibody)
FIG. 20 shows the immunohistochemical aspect of the tissue cross section of macrophages using anti-mouse Mac-3 antibody one week after CNT implantation. More positive cells were observed for Mac-3 in MWNT-II and CSNT than in SWNT and MWNT-I.

  The order of positive strength was as follows: MWNT-II = CSNT> MWNT-I> SWNT.

  FIG. 21 shows the immunohistochemical aspect using the Mac-3 antibody one month after implantation. It became clear that a larger number of positive cells were found around the SWNT material. The granulated tissue of SWNT was partially composed of macrophage components and no phagocytic cells were observed. However, there were fewer Mac-3 positive cells than SWNTs on granulospecies tissue in MWNT-I. Positive cells could be observed in a narrow space of self-aggregated bulk material. It was very interesting that phagocytic aspects were observed in MWNT-II (FIG. 21 (d)) and CSNT (FIG. 21 (e)).

(Comparison of granulation seed formation in the latter 3 months after implantation)
FIG. 22 shows the difference in granulation seed appearance and the arrangement of embedded CNTs in the later stage of this experimental prescription (3 months after implantation). Granulation seeds were expressed in all samples at 3 months after implantation, but the wall thickness was different depending on the substance attached. In particular, the wall thickness at SWNT was greater than the others. However, the degree of cell infiltration into massive CNTs increased in the order of CSNT, MWNT-II, and MWNT-I. In addition, there are loose boundaries between the granulation species, and the embedded material was found in particular MWNT-I. As a result of necropsy, no carbon material was found in the spleen, liver, kidney, lung, heart and cerebral cortex in all mice.

It is a transmission electron micrograph of the intermediate body of the fine carbon fiber obtained in the synthesis example. It is a transmission electron micrograph of the fine carbon fiber obtained in the synthesis example. It is an X-ray diffraction chart of the fine carbon fiber obtained in the synthesis example. It is a figure which shows typically the synthesizing | combining apparatus used in the synthesis example. It is a figure which shows typically the high temperature heat processing apparatus used in the synthesis example. It is the external appearance photograph (a) and micrograph (b) which show the dispersion state of the carbon fiber in the carbon fiber dispersion obtained in Example 1. FIG. 2 is a photograph showing a dispersion state of a carbon fiber dispersion obtained in Example 2. FIG. 2 is a photograph showing a dispersion state of a carbon fiber dispersion obtained in Comparative Example 1. It is the external appearance photograph (a) and micrograph (b) which show the dispersion state of the carbon fiber in the carbon fiber dispersion obtained in Example 3. FIG. 6 is a photograph showing the dispersion state of the carbon fiber dispersion obtained in Example 4. FIG. 6 is a photograph showing a dispersion state of a carbon fiber dispersion obtained in Comparative Example 2. It is a graph which shows the time-dependent change of the peripheral T cell after subcutaneous implantation of SWNT. It is a photograph which shows the state of the cell tissue from 1 week to 1 month after SWNT subcutaneous implantation. It is a graph which shows the time-dependent change of the peripheral T cell after the subcutaneous implantation of MWNT-I. It is a photograph which shows the state of the cell tissue from 1 week to 1 month after the subcutaneous implantation of MWNT-I. It is a graph which shows the time-dependent change of the peripheral T cell after the subcutaneous implantation of MWNT-II. It is a photograph which shows the state of the cell tissue from 1 week to 1 month after the subcutaneous implantation of MWNT-II. It is a graph which shows the time-dependent change of the peripheral T cell after the subcutaneous implantation of cap lamination CNT. It is a photograph which shows the state of the cell tissue from 1 week to 1 month after the subcutaneous implantation of the cap-stacked CNT. It is a photograph which shows the state of the tissue cross section of the macrophage using the anti-mouse Mac-3 antibody 1 week after CNT implantation. It is a photograph which shows the state of the tissue cross section of the macrophage using the anti-mouse Mac-3 antibody 1 week after CNT implantation. It is a photograph showing the difference in granulation seed appearance and the arrangement of embedded CNT 3 months after CNT implantation.

Claims (8)

  A carbon fiber dispersion wherein carbon fibers are dispersed in a polyol at a ratio of 0.001 to 30% by mass of the whole.   The carbon fiber dispersion according to claim 1, wherein the polyol is at least one selected from the group consisting of polyethylene glycol and propylene glycol.   The carbon fiber dispersion according to claim 1 or 2, wherein the carbon fiber is further diluted with an aqueous medium so that the carbon fiber is in a ratio of 0.001 to 30% by mass.   The carbon fiber dispersion according to any one of claims 1 to 3, wherein the carbon fiber dispersion is used for introduction into a living body.   A carbon fiber dispersion according to any one of claims 1 to 3 is introduced into a tissue or organ of an experimental animal, so that the carbon fiber is quantitatively placed in the body of the experimental animal. A method for evaluating the safety of carbon fibers, characterized by observing the state of the above.   Surgical procedure to incise the neck of the experimental animal to secure the airway, inject the carbon fiber dispersion into the lung of the experimental animal by injecting the carbon fiber dispersion into the lung, then suture the surgical site, 6. The method for evaluating the safety of carbon fiber according to claim 5, wherein the state of the experimental animal is subsequently observed.   After incising the skin of the experimental animal by surgical technique and injecting the carbon fiber dispersion subcutaneously, after implanting carbon fiber in the subcutaneous tissue of the experimental animal, the surgical site is sutured, and then the experimental animal The method for evaluating the safety of carbon fiber according to claim 5, wherein the state of the above is followed.   The method for evaluating the safety of carbon fiber according to any one of claims 5 to 7, wherein the experimental animal is a mouse.