JP6414949B2 - Self-curing chitosan-containing calcium phosphate composition, kit for producing the composition, and production method - Google Patents

Description

  The present invention relates to a self-curing chitosan-containing calcium phosphate composition, a kit for producing the composition, and a method for producing the composition.

  In recent years, in order to repair fractures or bone defects due to diseases or accidents, it has become common to use bone filling materials that are hydrated and cured in vivo. For example, in compression fracture of the vertebral body due to osteoporosis or bone tumor, percutaneous vertebroplasty is performed in which bone cement as a bone replacement material is percutaneously injected into the vertebral body with a puncture needle and the vertebral body is reconstructed. It has been broken.

  Currently, polymethyl methacrylate (PMMA) cement or calcium phosphate cement (CPC) is mainly used as bone cement. PMMA cement is excellent in curing speed and strength after curing, but lacks osteoconductivity and has poor affinity with bone tissue. Furthermore, PMMA cement has problems such as toxicity of unreacted monomers and damage of surrounding tissues due to generation of polymerization heat. On the other hand, CPC is inferior to PMMA cement in terms of curing speed and strength after curing, but is excellent in osteoconductivity because it is converted into bone-like hydroxyapatite in vivo. Since many calcium phosphate compounds, which are CPC materials, have the property of self-disappearing by being decomposed and absorbed in vivo and replaced with autologous bone, development of bone-replacement type CPC is also possible. Progressing. Moreover, CPC which improved the mechanical characteristic by mix | blending chitosan which is a biocompatible substance with CPC which is disclosed by the nonpatent literature 1 is also developed.

  In recent years, porous CPCs having a large number of minute voids called pores have been developed. In such a porous body, it is expected that osteoblasts, new bones, and the like enter the pores to form bone in the pores and to be integrated with the mother bone. In addition, since the surface area is increased by using a porous material, bone replacement in a living body is likely to occur. As such porous CPC, for example, a porous cured body described in Patent Document 1 can be mentioned. Patent Document 2 describes a filling material containing a substance that is mainly composed of a calcium phosphate compound such as hydroxyapatite and is decomposed and absorbed in a living body. After the filling material of Patent Document 2 is used in a living body, the material is made porous by being decomposed and absorbed in the living body.

JP 2004-269393 A JP 2004-049589 A

Sun L. et al., Fast setting calcium phosphate cement-chitosan composite: mechanical properties and dissolution rates, J. Biomater.Appl., Vol. 21, p. 299-315 (2007)

  However, in the porous CPC as described above, it usually takes about 2 to 3 months for the bone formation in the pores to improve the strength. Therefore, the porous cured body cannot be said to have sufficient initial strength immediately after transplantation into a living body. In percutaneous vertebroplasty, the CPC injected into the fracture site must withstand pressure, so it is important that the CPC has a high fracture energy, which is an indicator of toughness.

  Therefore, the present invention is self-curing that has excellent mechanical properties after curing, particularly toughness (magnification of fracture energy), and can obtain a cured body that is porous in vivo and replaced with autologous bone. An object of the present invention is to provide a type chitosan-containing calcium phosphate composition. The present invention also aims to provide a kit and a method for producing such a composition.

  The present inventor has produced CPC using tetracalcium phosphate, calcium hydrogen phosphate dihydrate, chitosan having an average molecular weight of 10,000 to 310,000, and malic acid aqueous solution as a hardening liquid as raw materials. It was found that the mechanical properties after curing were good. It was also found that the obtained CPC becomes porous in simulated body fluid or acetate buffer. Based on these findings, the present inventor has completed the present invention.

  Therefore, the present invention is a self-curing type obtained by mixing tetracalcium phosphate, calcium hydrogen phosphate dihydrate, chitosan having an average molecular weight of 10,000 or more and 310,000 or less, and malic acid aqueous solution as a hardening liquid. A chitosan-containing calcium phosphate composition is provided.

  The present invention also includes a self-curing chitosan containing a powder containing tetracalcium phosphate and calcium hydrogen phosphate dihydrate, and a hardening liquid containing chitosan and malic acid having an average molecular weight of 10,000 to 310,000. A kit for producing a calcium phosphate composition is provided.

  Furthermore, the present invention includes a step of mixing tetracalcium phosphate, calcium hydrogen phosphate dihydrate, chitosan having an average molecular weight of 10,000 to 310,000, and an aqueous malic acid solution as a hardening solution. A method for producing a chitosan-containing calcium phosphate composition is provided.

  According to the present invention, a self-curing chitosan-containing calcium phosphate composition excellent in mechanical properties after curing can be obtained. In addition, the cured product of the composition can be made porous in vivo and replaced with autologous bone.

2 is a graph showing the cure time of compositions containing chitosan at various concentrations. It is a graph which shows the compressive strength of the hardening body obtained from the composition containing chitosan in various density | concentrations. It is a graph which shows the fracture energy of the hardening body obtained from the composition containing chitosan in various density | concentrations. It is the electron micrograph of the hardening body after 1 day and 7 days after being immersed in ultrapure water. It is the electron micrograph of the hardening body after 1 day and 7 days after being immersed in Tris-HCl (pH 7.4). It is the electron micrograph of the hardening body after being immersed in the simulated body fluid (SBF) 7 days and 7 days. It is a graph which shows the calcium ion concentration in the acetic acid / sodium acetate buffer solution in which the hardening body was immersed. It is an electron micrograph of the hardening body before and behind being immersed in an acetic acid / sodium acetate buffer.

[Self-curing chitosan-containing calcium phosphate composition and method for producing the same]
Self-curing chitosan-containing calcium phosphate composition of the present invention (hereinafter also simply referred to as “composition”) is tetracalcium phosphate, calcium hydrogen phosphate dihydrate, chitosan, and malic acid aqueous solution as a curing solution. Is an amorphous mixture such as paste or slurry. The composition of the present invention immediately after mixing can be changed into an arbitrary shape or filled into a tube. And the composition of this invention self-hardens with time passage. The cured composition of the present invention is further cured by a hydration reaction in water or in vivo to become a cured product which is a kind of CPC. Therefore, the composition of the present invention can be suitably used as a bone substitute material.

  In the present invention, tetracalcium phosphate (TeCP) may be a commercially available product or one produced by itself. The TeCP production method itself is known, and any production method may be used. For example, it can be obtained as follows. Calcium carbonate powder and calcium hydrogen phosphate dihydrate powder are wet-mixed using water as a solvent, and the resulting mixture is fired at 1200-1700 ° C. for 3-6 hours. Then, by pulverizing the obtained fired product, TeCP powder can be obtained.

  In the embodiment of the present invention, the particle size of TeCP is not particularly limited, but is preferably less than 150 μm, more preferably less than 75 μm. A method for selecting particles having a desired particle diameter from TeCP powder is known per se, and any method can be used, but selection by a sieve is simple and preferable.

  In the composition of the present invention, by blending calcium hydrogen phosphate dihydrate (DCPD) together with TeCP, curing by the hydration reaction of the self-cured composition proceeds, and TeCP and DCPD are obtained in the obtained cured product. Reacts gradually with hydroxyapatite. In the embodiment of the present invention, the anhydrous calcium calcium phosphate (DCPA) may be used instead of DCPD. Commercially available DCPD and DCPA can be used.

  In the embodiment of the present invention, the particle size of DCPD is not particularly limited, but is preferably 0.2 to 10 μm, more preferably 0.5 to 5 μm. In addition, when the particle size of commercially available DCPD powder is not a preferred size, the particle size can be adjusted by pulverization by a known pulverization method such as wet pulverization.

  In the embodiment of the present invention, it is preferable to obtain a cement powder by dry mixing equimolar amounts of TeCP and DCPT. By using such cement powder, a uniform composition can be obtained.

  In the composition of the present invention, an aqueous malic acid solution is suitably used as the curing liquid from the viewpoint of shortening the curing time. The concentration of malic acid in the composition of the present invention is selected from the range of 1.0% by weight to 2.5% by weight, particularly 1.0% by weight to 2.0% by weight in consideration of the curing time and the compression strength of the cured product. It is desirable. When the malic acid concentration in the hardening liquid is 2.0% by weight or less, since chitosan described later cannot be dissolved in the hardening liquid, it is difficult to make the malic acid concentration in the composition 0.5% by weight or less. It is. On the other hand, when the malic acid concentration in the composition is larger than 2.5% by weight, the curing time of the composition is too short to be mixed.

  In the present invention, chitosan is used for the purpose of improving the curing rate of the self-curing calcium phosphate composition and the compressive strength of the cured product, and by eluting the cured product after injection or implantation into a living body, It is added for the purpose of making it porous. Chitosan is known as a natural polymer obtained by deacetylation of chitin contained in crustacean exoskeletons such as crabs and shrimps and fungal cell walls such as mushrooms with concentrated alkali. In the form, the origin of chitosan is not particularly limited.

  In the embodiment of the present invention, the form of chitosan is not particularly limited, and may be a powder form or a solution form. When preparing the composition of this invention using chitosan in the form of a solution, it is desirable for chitosan to melt | dissolve in malic acid aqueous solution as a hardening | curing liquid.

  In general, the molecular weight of chitosan is known to be proportional to the viscosity of the chitosan solution. The average molecular weight of chitosan used in the present invention is 10,000 to 310,000 as a molecular weight based on viscosity. In the present specification, the viscosity is measured with a B-type rotational viscometer at a solution temperature of 20 ° C. with respect to a chitosan solution obtained by dissolving chitosan in a 1% aqueous acetic acid solution to 1% by weight. Then, the intrinsic viscosity of chitosan is calculated based on the measured viscosity, and the molecular weight of chitosan is calculated from the Mark-Houwink-Sakurada equation.

  In the embodiment of the present invention, the degree of deacetylation of chitosan is not particularly limited, but is preferably 70 to 100%.

  In the present invention, a composition containing chitosan having a relatively medium molecular weight (190,000 or more and 310,000 or less) has a curing obtained by a hydration reaction than a composition containing chitosan having a relatively low molecular weight (less than 190,000). It tends to increase the destruction energy of the body, which is preferable.

  In an embodiment of the present invention, the concentration of chitosan in the composition ranges from 1.0% by weight to 10.0% by weight, particularly from 1.25% by weight to 3.75%, from the viewpoint of mechanical strength and porosity of the composition. It is desirable that the amount be in the range of weight percent or less. When the chitosan concentration in the hardening liquid is 20% by weight or more, the water content decreases, and the hardening liquid and the above-mentioned TeCP and DCPT cement powder cannot be mixed, so the chitosan concentration in the composition is 11.0%. It is difficult to make it more than% by weight.

  In the embodiment of the present invention, it is preferable to obtain a composition by mixing the above-mentioned cement powder of TeCP and DCPT with a hardening liquid containing malic acid and chitosan. Here, the powder-liquid ratio (powder (g) / liquid (g)) at the time of mixing cement powder and hardening liquid is 0.8-5 normally, Preferably it is 2-4, Most preferably, it is 3. is there. The mixing means is not particularly limited, and may be appropriately selected from known mixing means according to the mixing amount.

  After mixing the above cement powder and the curing liquid, the composition of the present invention usually completes self-curing in about 3 to 20 minutes (initial curing). The conditions of temperature and humidity at this time are not particularly different from general CPC preparation conditions, but are usually 20 to 40 ° C. and 50 to 100% humidity.

The obtained composition of the present invention can be used in a living body as a bone grafting material as described above. And the composition of this invention turns into a hardening body by hydration reaction in the living body. The cured product obtained from the composition of the present invention by a hydration reaction exhibits a compressive strength of at least 15 MPa, preferably at least 20 MPa, more preferably 25 MPa. Further, the cured product obtained from the composition of the present invention by a hydration reaction exhibits a fracture energy of at least 5.0 kJ / m ² , preferably at least 8.0 kJ / m ² , more preferably at least 10.0 kJ / m ² . Thereafter, chitosan contained in the cured body elutes in vivo to make the cured body porous. In addition, since this porosity occurs also by immersing a hardening body in physiological saline, after the hardening body is immersed in physiological saline, it can confirm with a scanning electron microscope etc. it can.

[Kit for producing self-curing calcium phosphate composition]
A kit for producing the self-curing calcium phosphate composition of the present invention (hereinafter also simply referred to as “kit”) includes a powder containing tetracalcium phosphate and calcium hydrogen phosphate dihydrate, and an average molecular weight of 10,000. A hardener containing ˜310,000 chitosan and malic acid.

  In the embodiment of the present invention, the powder containing two groups of TeCP and DCPD having different particle sizes is preferably a cement powder obtained by dry-mixing equimolar amounts of TeCP and DCPT described above. The details of TeCP and DCPT are the same as those described in the description of the composition of the present invention.

  In the embodiment of the present invention, the hardening liquid containing chitosan and malic acid is preferably the above-described malic acid aqueous solution in which chitosan is dissolved. The details of each of chitosan and malic acid are the same as those described in the description of the composition of the present invention. Further, the malic acid concentration and chitosan concentration in the curable liquid are the same as those described in the description of the composition of the present invention.

  In the embodiment of the present invention, the kit is preferably a two-reagent kit in which a powder containing TeCP and DCPD and a hardening liquid containing chitosan and malic acid are contained in separate containers. The shape of such a container is not particularly limited, and for example, the powder containing TeCP and DCPD is stored in a kneading injector that can be mixed with the curable liquid, and the curable liquid is also stored in the injector. In this case, the kit of the present invention may further include an injection needle. Moreover, the kit of this invention may further contain the water for adjusting the viscosity of the composition obtained.

  EXAMPLES The present invention will be described in detail below by examples, but the present invention is not limited to these examples.

<Example 1>
In this example, the curing time of the composition of the present invention and the mechanical properties of the cured product obtained from the composition were examined.
(1) Materials: Tetracalcium phosphate (TeCP)
Calcium carbonate (11.032 g (0.110 mol), Wako Pure Chemical Industries, Ltd.), calcium hydrogen phosphate dihydrate (18.968 g (0.110 mol), Wako Pure Chemical Industries, Ltd.), and ultrapure water (180 mL ) Together with zirconia balls (200 g, manufactured by ASONE CORPORATION) in a pot mill HD-A-4 (manufactured by ASONE CORPORATION) and wet-mixed at room temperature for 24 hours at a rotation speed of 110 rpm. The obtained slurry was placed on filter paper 5C (manufactured by Kiriyama Seisakusho) and suction filtered with an aspirator A-3S (manufactured by EYELA). The filtrate was put in a petri dish and dried at 50 ° C. for 24 hours with a constant temperature dryer DOV-450P (manufactured by ASONE Corporation). The dried product is pulverized, and the obtained powder is placed in a hot press machine mold 1-6002-18 (manufactured by ASONE Co., Ltd.) having an inner diameter of 14 mm, and 1,100 kgf / cm ² using a press (manufactured by Jusco Engineering). A cylindrical sample having a height of about 12 mm was obtained. The obtained sample was placed in an alumina boat SSA-H2B (manufactured by Nikkato Co., Ltd.) and baked in the atmosphere at 1500 ° C. for 5 hours using an electric furnace (manufactured by Marusho Denki Co., Ltd.). The temperature raising / lowering rate was 10 ° C / min. After firing, the sample was taken out of the electric furnace, pulverized with a super hard steel mortar (WD type, manufactured by Ito Seisakusho), and further finely pulverized with an agate mortar to obtain TeCP powder. The obtained TeCP powder was classified using a sieve shaker MVS-1 (manufactured by ASONE Co., Ltd.) to obtain a TeCP powder having an average particle size of 20.43 μm. The frequency of the sieve shaker was 10 rps, and the sieve opening was 75 μm.

・ Calcium hydrogen phosphate dihydrate (DCPD)
Calcium hydrogen phosphate dihydrate (15.00 g, Wako Pure Chemical Industries, Ltd.) and ultrapure water (180 mL) together with zirconia balls (500 g, manufactured by As One Corporation), Pot Mill HD-A-4 (As One And was wet-ground for 48 hours at a rotation speed of 110 rpm at room temperature. The obtained slurry was placed on filter paper 5C (manufactured by Kiriyama Seisakusho) and suction filtered with an aspirator A-3S (manufactured by EYELA). The filtrate was put in a petri dish and dried at 50 ° C. for 24 hours with a constant temperature dryer DOV-450P (manufactured by ASONE Corporation). The dried product was pulverized to obtain a DCPD powder having an average particle size of 1.76 μm.

・ Curing solution As the curing solution, 10% by weight of malic acid (Wako Pure Chemical Industries, Ltd.) and 5, 10, 15 or 20 of chitosan (average molecular weight 50,000 to 190,000, deacetylation degree 81.3%, ARDICRH Chemistry) An aqueous solution containing 10% by weight of malic acid and an aqueous solution containing 5, 10, 15 or 20% by weight of malic acid and chitosan (average molecular weight 190,000-310,000, degree of deacetylation about 78.8%, ARDRICH Chemistry) were used. . As a control, a hardening solution containing only malic acid (10% by weight) was used.

(2) Preparation of self-curing calcium phosphate composition and cured product thereof
Put TeCP powder (6.80 g (0.186 mol)) and DCPD powder (3.20 g (0.186 mol)) in a 50 ml threaded test tube (manufactured by ASONE Corporation), and shaker MALTI SHAKER MS- It was installed at 300 (manufactured by ASONE Co., Ltd.) and mixed with shaking at a rotational speed of 1300 rpm for 100 minutes. The obtained mixture was further frictionally mixed in an agate mortar for 5 minutes to obtain cement powder. The particle size distribution of the cement powder was examined with a laser diffraction / scattering type particle size distribution analyzer LA-920 (manufactured by Horiba, Ltd.). Cement powder and hardened liquid are mixed for 90 seconds so that the powder-liquid ratio (powder (g) / liquid (g)) is 3, self-setting calcium phosphate composition (malic acid 2.5 wt%, chitosan 0, 1.25, 2.5 or 3.75% by weight) was obtained. However, when a curable liquid containing 20% by weight of chitosan was used, it was not possible to mix due to the low water content. A part of the composition obtained using a curable liquid containing 2.5 to 15% by weight of chitosan of each molecular weight was taken, and this was used as a sample for measurement of the curing time. The remaining composition was poured into a Teflon split mold (inner diameter 6 mm, height 12 mm) and allowed to stand in an environment of 37 ° C. and 100% humidity for 1 hour to obtain an initial cured product. Then, the initial cured body was taken out of the mold and immersed in ultrapure water for 24 hours to be hydrated and cured to obtain a cured body. The obtained cured body was subjected to measurement of mechanical properties as a sample.

(3) Sample analysis
(3-1) Measurement of curing time Each sample was placed in a Teflon mold (inner diameter: 10 mm, height: 5 mm) within 90 seconds from the start of mixing of the above cement powder and the curing liquid. Then, after 30 minutes from the start of mixing, the needle (300 g, tip cross-sectional area 2 mm ² ) of the Vicat needle tester (manufactured by Nippon Mec Co., Ltd.) is dropped on the sample surface until no indentation remains on the surface. The time was recorded. In addition, except at the time of the test by a Vicat needle tester, each sample was stored at 37 ° C. in a closed container equipped with a sponge containing water at the bottom. In this example, the humidity in the sealed container was regarded as 100%. The measurement was performed three times for each sample, and the average value and standard error of the curing time were calculated based on the measured value (minute). The results are shown in FIG. In FIG. 1, the horizontal axis represents the chitosan concentration (% by weight) in the composition, and the vertical axis represents the curing time (minutes).

  From FIG. 1, it was found that a composition containing chitosan having a molecular weight of 190,000 or more and 310,000 or less has a longer curing time than a composition containing chitosan having a molecular weight of 50,000 or more and less than 190,000.

(3-2) Measurement of Compressive Strength and Fracture Energy A compression test of each sample was performed according to JIS standard (JIS T6602) using a universal testing machine AG-10 (manufactured by Shimadzu Corporation). In this test, a 5 kN load cell was used, and the test speed was 0.50 mm / sec. The test was performed six times for each sample, and the average value and standard error of the compressive strength were calculated based on the measured values. In addition, a stress-strain curve was created based on the measured values, and the fracture energy was calculated from the area surrounded by the curve. The results are shown in FIGS. In FIG. 2, the horizontal axis indicates the chitosan concentration (% by weight) in the composition, and the vertical axis indicates the compressive strength (MPa). In FIG. 3, the horizontal axis represents the chitosan concentration (% by weight) in the composition, and the vertical axis represents the fracture energy (kJm ² ).

(4) Results and Discussion From FIGS. 2 and 3, it was found that the inclusion of chitosan in the calcium phosphate composition increases the compressive strength and fracture energy of the cured product obtained by the hydration reaction. From FIG. 2, it was found that the cured product obtained from the composition of the present invention had a compressive strength of at least 17 MPa in this example. Further, FIG. 3 shows that the breaking energy of the composition containing chitosan having a molecular weight of 190,000 or more and 310,000 or less is larger than that of the composition containing chitosan having a molecular weight of 50,000 or more and less than 190,000.

<Example 2>
In this example, whether or not the cured body became porous was tested in ultrapure water, Tris-HCl (pH 7.4) or simulated body fluid (SBF).

(1) Materials, TeCP and DCPD
TeCP and DCPD obtained in the same manner as in Example 1 were used.
・ Curing solution As the curing solution, an aqueous solution containing 10% by weight of malic acid (Wako Pure Chemical Industries, Ltd.) and 10% by weight of chitosan (average molecular weight 190,000-310,000, degree of deacetylation about 78.8%, ARDICRH Chemistry) Using.
-Simulated body fluid (SBF)
SBF was prepared as described in Oyane A. et al., J. Biomed. Mater. Res., 65A, 188-195 (2003).

(2) Preparation of self-curing calcium phosphate composition and cured body Cement powder was prepared from TeCP and DCPD in the same manner as in Example 1. Using the obtained cement powder and the above-mentioned hardening liquid, the mixture is mixed for 90 seconds so that the powder-liquid ratio (powder (g) / liquid (g)) is 3, and the self-hardening calcium phosphate composition (malic acid 2.5% by weight and 2.5% by weight of chitosan). This composition was poured into a Teflon mold (inner diameter 10 mm, height 2 mm) and allowed to stand in an environment of 37 ° C. and 100% humidity for 1 hour to obtain an initial cured product.

(3) Analysis of sample The obtained cured product was immersed in ultrapure water, Tris-HCl (pH 7.4) or SBF at 37 ° C, and the cured product was subjected to field emission scanning after 1 day and 7 days later. Observation was performed with an electron microscope JSM-6500F (manufactured by JEOL Ltd.). The obtained electron micrographs are shown in FIGS. 4 to 6 show that the cured product obtained from the composition of the present invention becomes porous even when immersed in any of ultrapure water, Tris-HCl (pH 7.4), and SBF. It was done.

<Example 3>
In this example, the relationship between the calcium ion elution amount and the weight loss amount of the cured body was examined in a pH 5.5 buffer solution that is the same as the environment in the presence of osteoclasts in vivo.

(1) Materials, TeCP and DCPD
TeCP and DCPD obtained in the same manner as in Example 1 were used.
・ Curing solution As the curing solution 1, an aqueous solution containing 10% by weight of malic acid (Wako Pure Chemical Industries, Ltd.) and 10% by weight of chitosan (average molecular weight 190,000-310,000, degree of deacetylation about 78.8%, ARDICRH Chemistry) Was used. Further, as hardening solution 2, an aqueous solution containing 10% by weight of malic acid (Wako Pure Chemical Industries, Ltd.) and 18% by weight of chitosan (average molecular weight 190,000-310,000, degree of deacetylation about 78.8%, ARDICRH Chemistry) Using. As a control, ultrapure water was used as a curable liquid.

Acetic acid / sodium acetate buffer solution Acetic acid (Wako Pure Chemical Industries, Ltd.) was added to ultrapure water to prepare an acetic acid aqueous solution (0.08 mol / L). Also, sodium acetate (Wako Pure Chemical Industries, Ltd.) was dissolved in ultrapure water to prepare an aqueous sodium acetate solution (0.08 mol / L). The prepared acetic acid aqueous solution (200 mL) and sodium acetate aqueous solution (1400 mL) were mixed to prepare an acetic acid / sodium acetate buffer. The pH of this acetic acid / sodium acetate buffer was 5.5.

(2) Preparation of self-curing calcium phosphate composition and cured body Cement powder was prepared from TeCP and DCPD in the same manner as in Example 1. Using the obtained cement powder and the above-mentioned hardening liquid 1, the mixture is mixed for 90 seconds so that the powder-liquid ratio (powder (g) / liquid (g)) is 3, and the self-hardening calcium phosphate composition (apple Acid 2.5% by weight and chitosan 2.5% by weight: hereinafter referred to as “CPCch2.5”). Further, using the above cement powder and the above hardening liquid 2, the mixture is mixed for 90 seconds so that the powder-liquid ratio (powder (g) / liquid (g)) is 0.8, and the self-curing calcium phosphate composition ( Containing 2.5% by weight of malic acid and 10.0% by weight of chitosan: hereinafter referred to as “CPCch10”). Further, using the above cement powder and the above-described ultrapure water, the mixture is mixed for 90 seconds so that the powder / liquid ratio (powder (g) / liquid (g)) is 3, and the self-curing calcium phosphate composition ( Malic acid and chitosan free: hereinafter referred to as “CPCcont”). These compositions were poured into a Teflon mold (inner diameter 10 mm, height 2 mm) and allowed to stand in an environment of 37 ° C. and 100% humidity for 1 hour to obtain an initial cured product. The obtained initial cured product was immersed in physiological saline (37 ° C.) for 7 days to obtain a cured product, which was dried at 50 ° C. for 24 hours. A dried cured body was used as a sample. In addition, before measuring the calcium ion elution amount, the weight of each sample was measured.

(3) Sample analysis
(3-1) Measurement of calcium ion elution amount Immerse the cured body in acetic acid / sodium acetate buffer solution (100 mL, 37 ° C), and measure the amount of calcium ions eluted from the cured body in the buffer solution. Measurements were made over time up to 300 minutes. More specifically, measurement was performed every 2 minutes until 20 minutes after immersion, measurement was performed every 15 minutes from 20 minutes to 150 minutes after immersion, and measurement was performed every 30 minutes from 150 minutes to 300 minutes after immersion. For the measurement, a calcium ion electrode (manufactured by Metrohm) was used. The results are shown in FIG.

(3-2) Measurement of weight loss Each cured product after being immersed in an acetic acid / sodium acetate buffer solution for 300 minutes was dried at 50 ° C. for 24 hours. Then, the weight of each dried cured body was measured. And the weight reduction amount (weight before immersion-weight after immersion) and the difference (weight reduction amount-calcium ion elution amount) of both were computed. An electron micrograph obtained by photographing CPCch10 before immersion in acetic acid / sodium acetate buffer and after immersion for 300 minutes with a field emission scanning electron microscope JSM-6500F (manufactured by JEOL Ltd.) is shown in FIG. Shown in

(4) Results and Discussion From FIG. 7, the calcium ion concentration in the acetic acid / sodium acetate buffer solution, that is, the calcium elution amount from the cured product, CPCcont becomes the highest with time, and then CPCch2.5 becomes higher. CPCch10 was found to be the lowest. On the other hand, the weight loss of each cured product was highest for CPCch10, then higher for CPCch2.5, and lowest for CPCcont. These facts suggest that chitosan is preferentially eluted in an acetic acid / sodium acetate buffer in a cured product obtained from a composition containing chitosan. Moreover, the electron micrograph of FIG. 8 showed that the cured product containing chitosan was porous in an acetic acid / sodium acetate buffer.

Claims (5)

A self-curing chitosan-containing calcium phosphate composition comprising tetracalcium phosphate, calcium hydrogen phosphate dihydrate, chitosan having an average molecular weight of 10,000 to 310,000, and malic acid aqueous solution as a hardening liquid. There,
The concentration of chitosan in the composition is 1.0 wt% or more and 3.75 wt% or less, and the concentration of malic acid in the composition is 1.0 wt% or more and less than 2.5 wt%,
Self-curing chitosan-containing calcium phosphate composition.   The self-curing chitosan-containing calcium phosphate composition according to claim 1, wherein the average molecular weight of chitosan is 190,000 or more and 310,000 or less.   The self-hardening chitosan-containing calcium phosphate composition according to claim 1 or 2, wherein chitosan is dissolved in an aqueous malic acid solution as a hardening liquid.   The powder containing tetracalcium phosphate and calcium hydrogenphosphate dihydrate, and the hardening liquid containing chitosan and malic acid having an average molecular weight of 10,000 to 310,000. A kit for producing a self-curing chitosan-containing calcium phosphate composition.   4. The method according to claim 1, comprising a step of mixing tetracalcium phosphate, calcium hydrogenphosphate dihydrate, chitosan having an average molecular weight of 10,000 to 310,000, and malic acid aqueous solution as a hardening solution. The manufacturing method of self-hardening type chitosan containing calcium phosphate composition as described in an item.