WO2019077806A1

WO2019077806A1 - Bone graft material and method for manufacturing bone graft material

Info

Publication number: WO2019077806A1
Application number: PCT/JP2018/024191
Authority: WO
Inventors: 怜本島; 山田　真也
Original assignee: オリンパステルモバイオマテリアル株式会社
Priority date: 2017-10-20
Filing date: 2018-06-26
Publication date: 2019-04-25

Abstract

The purpose of the present invention is to further improve osteogenic properties and shape maintainability. The present invention provides a bone graft material that comprises calcium phosphate and a biodegradable polymer in a mass ratio of 4:1 or more and less than 10:1, wherein the calcium phosphate comprises a plurality of particles that have different particle sizes with the most frequent diameter of the particles being 20 μm or less, and the biodegradable polymer is crosslinked.

Description

Bone filling material and method for producing bone filling material

The present invention relates to a bone graft material and a method of manufacturing the bone graft material.

Conventionally, as a bioabsorbable bone filling material having high bone conductivity, organic materials such as hydroxyapatite (HA) which is an inorganic material and calcium phosphate based ceramics typified by β-tricalcium phosphate (β-TCP) are used. Composite materials produced by mixing collagen, which is a material, are widely used (see, for example, Patent Documents 1 and 2).

Unexamined-Japanese-Patent No. 2014-76387 specification U.S. Patent Application No. 2011/0017166

Patent Document 1 describes a bone graft material produced by mixing hydroxyapatite (HA) and collagen. However, since the bone filling material of Patent Document 1 has a mass ratio of hydroxyapatite (HA) to collagen of 1:10 to 5: 1 (w / w), the mass of hydroxyapatite with respect to collagen is small. There is a problem that a sufficient new bone formation effect can not be obtained. Patent Document 2 describes a bone graft material produced by mixing β-type tricalcium phosphate (β-TCP) and collagen, but the particle size of β-type tricalcium phosphate granules used is As large as 100 μm to 300 μm, when dispersed on a porous collagen matrix, as shown in FIG. 12, localized in a non-uniform manner on the collagen matrix, and the scaffolding environment for bone formation worsens. There is a problem that the bone conduction ability is inhibited.

The present invention has been made in view of the above-described circumstances, and it is an object of the present invention to provide a bone grafting material and a method of producing the bone grafting material, which can further enhance bone formation and shape maintenance.

In order to achieve the above object, the present invention provides the following means.
One aspect of the present invention comprises calcium phosphate and a biodegradable polymer in a mass ratio of 4: 1 to 10: 1, wherein the calcium phosphate comprises a plurality of particles having different particle sizes, and the mode of the particles It is a porous bone grafting material having a diameter of 20 μm or less and the biodegradable polymer being crosslinked.

According to the above aspect, since the mass ratio of calcium phosphate to the biodegradable polymer is appropriately defined as 4: 1 or more and less than 10: 1, it is sufficient while having sufficient flexibility and shape recovery. Bone grafting material capable of exerting new bone formation ability can be produced. Therefore, the bone defect portion can be filled with the bone filling material without gaps, and a sufficient new bone formation effect can be obtained. Since the mode diameter of calcium phosphate is 20 μm or less, the calcium phosphate can be uniformly and densely dispersed throughout the biodegradable polymer matrix, and the scaffolding environment for new bone formation can be improved. As a result, the new bone forming ability can be further improved.
The “mode diameter” is the maximum value of volume% or number% with respect to the particle diameter in the particle size distribution of particles, and is also called mode diameter.

In the above aspect, the pore diameter of the bone graft material may be larger than 20 μm.
The porosity of the bone graft material may be 50% or more.
In this way, the pore diameter of the bone filling material necessary for the invasion of tissues such as cells and blood vessels is secured, and a space capable of circulating blood and body fluid necessary for cell growth and formation of bone tissue. Can be secured. As a result, new bone formation can be further promoted.

In the above aspect, the calcium phosphate may be β-tricalcium phosphate (β-TCP).
In this way, a bone graft material having excellent bone conductivity and high bioresorbability can be produced.

In the above aspect, the biodegradable polymer may include at least collagen.
In this way, a flexible bone substitute can be easily generated.

In the above aspect, the crosslinking may be chemical crosslinking.
In this way, the elasticity, flexibility and shape recovery of the bone filling material produced can be improved, and a space for cells and tissues necessary for new bone formation to be invaded can be maintained. it can. The stability of the scaffolding environment for new bone formation can be improved.

In the above aspect, the chemical crosslinking agent used for the chemical crosslinking may include at least one of epoxide and carbodiimide.
By doing this, it is possible to obtain a bone graft material having high shape recovery even in a wet state and high biosafety.

Another aspect of the present invention comprises the steps of: calcining a precursor containing calcium phosphate to form a calcium phosphate fine powder; mixing the calcium phosphate fine powder with a biodegradable polymer solution and stirring; and the stirring step A cross-linking step of forming a three-dimensional biodegradable polymer / calcium phosphate composite suspension by adding a cross-linking agent to the resulting product to form a three-dimensional biodegradable polymer / calcium phosphate composite suspension, and the biodegradable material produced by the cross-linking step Lyophilizing polymer / calcium phosphate composite suspension, unreacted crosslinker and reaction by-product from the sponge-like biodegradable polymer / calcium phosphate composite porous body obtained by the lyophilization step And washing the biodegradable polymer / calcium phosphate composite porous material washed and removed. And B. producing a bone graft material, wherein a mass mixing ratio of the calcium phosphate and the biodegradable polymer is 4: 1 or more and less than 10: 1, and the calcium phosphate has a plurality of different particle sizes. And the mode diameter of the particles is 20 μm or less.

According to the above aspect, since the mass ratio of calcium phosphate to the biodegradable polymer is appropriately defined as 4: 1 or more and less than 10: 1, it is sufficient while having sufficient flexibility and shape recovery. Bone grafting material that exerts a new bone forming ability can be produced. Since the mode diameter of the calcium phosphate particles is 20 μm or less, the calcium phosphate particles can be uniformly and densely dispersed throughout the biodegradable polymer matrix, and the scaffolding environment for new bone formation can be improved. As a result, the new bone forming ability can be further improved.

In the above aspect, the pore diameter of the bone graft material is preferably larger than 20 μm.
The porosity of the bone graft material is preferably 50% or more.
In this way, the pore diameter of the bone filling material necessary for the invasion of tissues such as cells and blood vessels can be secured, and the circulation of blood, body fluid, and oxygen necessary for cell growth and formation of bone tissue is possible. Space can be secured. As a result, new bone formation can be further promoted.

In the above aspect, the calcium phosphate is preferably β-tricalcium phosphate (β-TCP).
In this way, a bone graft material having excellent bone conductivity and high bioresorbability can be produced.

In the above aspect, the biodegradable polymer preferably contains at least collagen.
By doing so, a flexible bone graft material can be easily produced.

In the above embodiment, the crosslinking is preferably chemical crosslinking.
In this way, the elasticity, flexibility and shape recovery of the bone filling material produced can be improved, and a space for cells and tissues necessary for new bone formation to be invaded can be maintained. it can. Also, the stability of the scaffolding environment for new bone formation can be improved.

In the above aspect, it is preferable that the chemical crosslinking agent used for the chemical crosslinking contains at least one of an epoxide and a carbodiimide.
By doing this, it is possible to obtain a bone graft material having high shape recovery even in a wet state and high biosafety.

ADVANTAGE OF THE INVENTION According to this invention, it is effective in the ability to provide the manufacturing method of the bone grafting material which can further improve new bone formation property and shape maintenance property, and a bone grafting material.

It is sectional drawing of the bone grafting material which concerns on one Embodiment of this invention. It is a figure which shows the production | generation process of the bone grafting material of FIG. It is a scanning electron micrograph of the bone graft material cross section of FIG. It is a scanning electron micrograph of a bone graft material cross section which has a porosity of 50% or more and an average pore diameter of 50 μm or more, and is a low magnification photograph of 100 times. It is a scanning electron micrograph of a bone graft material cross section having a porosity of 50% or more and an average pore diameter of 50 μm or more, which is a high magnification photograph of 300 times. FIG. 2 is a flowchart showing a method of producing the bone graft material of FIG. 1, wherein (a) is a β-TCP fine powder production step, (b) is a heat denatured collagen production step, and (c) is a bone grafting material production step Respectively. It is a graph which shows the experimental result of Example 1, and is a graph which evaluated shape restoration nature of a bone grafting material. It is a photograph of the filling part of the rat in which the bone filling material was filled in the bone defect part of the skull. In Example 2, it is a μCT image which shows an example of a score used for determination of bone conductivity, and it is shown that the transplanted bone graft material has united in the full range. In Example 2, a μCT image showing an example of a score used for determination of bone conductivity, which is shown to be partially fused. In Example 2, it is a μCT image showing an example of a score used for determination of bone conductivity, and it is shown that healing was not confirmed. In Example 2, it is a μCT image which shows an example of a score used for determination of bone approachability, and it is shown that the transplanted bone substitute material has entered into the central part of a bone defect. In Example 2, it is a μCT image showing an example of a score used for determination of bone approachability, and it is shown that new bone is formed only at the peripheral portion of a bone defect. In Example 2, it is a μCT image showing an example of a score used for determination of bone penetration, and it is shown that bone penetration was not confirmed. It is a μCT image which is an experimental result of six weeks after transplanting the bone grafting material which concerns on Example 2 to the skull of a rat, and shows the state of the bone regeneration degree of sample AK. 5 is a graph showing experimental results of Example 2. It is a figure which shows the production | generation process of the bone grafting material which concerns on a prior art. It is a scanning electron micrograph of the cross section of the bone graft material produced in FIG. Low and high magnification photographs of bone filler cross sections of Samples AJ.

Below, the bone grafting material concerning one embodiment of the present invention is explained.

Below, the bone grafting material concerning one embodiment of the present invention is explained with reference to drawings.
The bone grafting material 1 according to the present embodiment, as shown in FIGS. 1 and 2, includes a collagen matrix 2 mainly composed of collagen which is a biodegradable polymer, and β-TCP (β-type phosphate 3) It is a collagen / β-TCP composite material obtained by mixing it with a fine powder of calcium 3). After being implanted in a bone defect, β-TCP3 is designed to be gradually absorbed into the body by the action of surrounding bone tissue and to be replaced with autologous bone.

The collagen matrix 2 is crosslinked by the crosslinking agent 5. Thereby, the collagen molecules are cross-linked to form a three-dimensional structure, whereby the shape can be maintained for a certain period in the living body while having a certain elasticity. In the present embodiment, the crosslinking treatment is particularly preferably crosslinking by chemical crosslinking.
As the chemical crosslinking agent, crosslinking agents such as epoxides and carbodiimides are particularly preferable in consideration of the easiness of crosslinking reaction and the biocompatibility of the obtained bone filler 1, but the invention is not limited thereto.

The fine powder of β-TCP3 contains a plurality of particles having different particle sizes, and the mode diameter is 20 μm or less. When the mode diameter of the fine powder particles of β-TCP3 is larger than 20 μm, the action to promote bone formation may be reduced. The fine powder of β-TCP3 is complexed in such a manner as to be taken into the pore walls of collagen matrix 2.

As shown in FIG. 1, FIG. 3 and FIGS. 4A and 4B, a plurality of pores 4 having an average diameter of 50 μm or more are formed in the bone graft material 1. The scanning electron microscope (SEM) image of the bone grafting material 1 cross section in the case where porosity is 50% or more and average pore diameter is 50 micrometers or more is shown by FIG. 4A and 4B. FIG. 4A shows an SEM image of the bone grafting material 1 observed at a low magnification of 100 ×, and FIG. 4B shows an SEM image of the bone grafting material 1 observed at a high magnification of 300 ×. By forming a plurality of such pores 4 in the bone filling material 1, it is possible to promote the invasion of tissues such as cells and blood vessels, enable circulation of blood and body fluid, and efficiently promote bone formation. It is supposed to be.

The bone grafting material 1 according to the present embodiment is formed such that the mass ratio of β-TCP 3 to collagen matrix 2 is 4: 1 or more and less than 10: 1. Therefore, it has become possible to exert sufficient bone formation ability while having sufficient flexibility and shape recovery.

In this embodiment, as calcium phosphate, one using β-TCP, which can be stably present in a living body for a long period of time and is particularly excellent as a biomaterial, has been described, but the present invention is not limited thereto. It may be compatible, that is, applicable as a biomaterial. For example, as a calcium phosphate compound, apatites such as hydroxyapatite, dicalcium phosphate, tetracalcium phosphate, octacalcium phosphate etc. may be mentioned, and one or more of them may be used in combination it can.

In this embodiment, although what used collagen as biodegradable polymer was explained, it is good also as replacing with this, using pectin, sodium alginate, gelatin, etc., and it is not restricted to this.

The bone grafting material 1 according to this embodiment configured as described above is manufactured by the following manufacturing method.
In the method of producing the bone graft material 1 according to the present embodiment, as shown in FIGS. 5A to 5C, a β-TCP fine powder production process SA for producing a fine powder of β-TCP 3 and heat denaturation The heat-denatured collagen production process SB for producing collagen, the β-TCP fine powder production process SA and the heat-denatured collagen production process SA1 And a bone grafting material manufacturing step SC for manufacturing the bone grafting material 1.

As shown in FIG. 5 (a), the β-TCP fine powder production process SA includes a synthesis step SA1 of synthesizing a precursor of β-TCP3 and a step of baking the synthesized precursor of β-TCP3. A baking step SA2 for producing a TCP aggregate, and a grinding step SA3 for grinding the produced β-TCP3 aggregate.
The synthesis step SA1 is carried out, for example, by drying a slurry containing a precursor of β-TCP3 synthesized from a calcium supplying material and a phosphoric acid supplying material. A precursor of β-TCP3 is obtained by this synthesis step SA1.
The firing step SA2 is performed by firing the precursor of β-TCP3 obtained by step SA1. Thereby, an aggregate of β-TCP3 is obtained.
The crushing step SA3 is performed, for example, by crushing the β-TCP3 aggregate obtained in step SA2 sufficiently finely. Thereby, a fine powder of β-TCP3 is obtained. The method of grinding is not particularly limited.

As shown in FIG. 5 (b), the heat-denatured collagen production process SB is obtained by step SB1 of dissolving atelocollagen atelolated by pepsin treatment in an acidic solvent to prepare an atelocollagen acidic solution, and step SB1. Prepare an acidic solution of atelocollagen by lyophilization, and add an appropriate amount of phosphate buffer solution to the collagen sponge obtained by the drying step SB2, and heat treatment to denature the collagen to obtain heat-denatured collagen And degenerative step SB3.

As shown in FIG. 5 (c), the bone filling material production process SC includes the fine powder of β-TCP3 obtained by the β-TCP fine powder production process SA and the atelocollagen obtained by the heat-denatured collagen production process SB. A mixing step SC1 of mixing and stirring with an acidic solution, a collagen / β-TCP composite gel forming step SC2 of obtaining a collagen / β-TCP composite gel from the mixed mixture, and a collagen / β-TCP composite Stir the gel to break up the gel and suspend the collagen / β-TCP composite gel stirring step SC3 and the product obtained by stirring the collagen / β-TCP composite gel by adding phosphate buffer Collagen / β-TCP composite suspension preparation step of preparing collagen / β-TCP composite suspension step SC4 and heat denaturation to collagen / β-TCP composite suspension Heat denatured collagen obtained in the heat denaturation step SB3 to obtain lagen Heat addition denatured collagen addition step SC5, and crosslinking step SC6 to carry out the crosslinking treatment by adding the crosslinking agent 5 to the collagen / β-TCP composite material suspension And lyophilization of the cross-linked collagen / β-TCP composite suspension by the lyophilization step of collagen / β-TCP composite suspension lyophilization step SC7 and the collagen / β-TCP composite suspension lyophilization step SC7 Freeze the collagen / β-TCP composite after the unreacted crosslinker 5 is removed by the washing step SC8 for washing the unreacted crosslinking agent 5 from the obtained collagen / β-TCP composite and the washing step SC8 Process the drying step SC9 to dry and the dried sponge-like collagen / β-TCP composite to make it elastic in any shape And processing step SC10 for producing the bone grafting material 1 having.

The bone graft material 1 according to this embodiment manufactured in this manner is used for treatment of a bone defect by being implanted in a bone defect in a living body. The transplanted bone filling material 1 serves as a scaffold when osteoblasts form new bone, and new bone is formed in the bone defect part with the passage of time. Thereby, the bone defect can be cured.

According to the present embodiment, by setting the mass ratio of the fine powder of β-TCP 3 to the collagen matrix 2 to be 4: 1 to 10: 1, the elasticity is improved compared to the conventionally used bone graft material. Can rapidly and sufficiently form new bone while achieving sufficient flexibility and shape recovery. Collagen of β-TCP3 fine powder with calcium phosphate particles having a mode diameter of 20 μm or less while securing pores that allow blood and osteoblasts to penetrate into the bone filling material 1 transplanted without gaps in the bone defect part without gaps By uniformly and densely dispersing all over the matrix 2, the fine powder of β-TCP 3 can be in close contact with the bone defect around the transplanted part. As a result, by inducing osteoblasts necessary for new bone formation from the periphery of the bone defect into the bone filling material 1, it is possible to promote the formation of new bone.

Next, Examples 1 to 5 of the embodiment of the present invention described above will be described below with reference to the drawings.
Example 1
(Shape recovery evaluation)
Samples of a total of four bone grafting materials 1 in which the blending mass ratio of β-TCP and collagen was changed were prepared, and the shape recovery of each of the prepared samples was evaluated by the following procedure.
First, the following four samples represented by xBTy were prepared, where x is a mass ratio of β-TCP when collagen is 1, and y is a mode diameter of β-TCP fine powder particles. The most frequent diameter y of the β-TCP fine powder particles was all prepared to be less than 10 μm.
Sample 1: 4BT10 (β-TCP: collagen = 4: 1 (w / w))
Sample 2: 9BT10 (β-TCP: collagen = 9: 1 (w / w))
Sample 3: 13BT 10 (β-TCP: collagen = 13: 1 (w / w))
Sample 4: 16BT10 (β-TCP: collagen = 16: 1 (w / w))
Samples 1-4 were all processed into 1 cm ³ cubes.

Next, the shape recoverability of the prepared samples 1 to 4 was evaluated by the following experiment.
First, RO water (Reverse Osmosis water) was dropped to each of the samples 1 to 4 until the maximum water absorption capacity was exceeded and water absorption became impossible. Next, samples 1 to 4 which had been sufficiently absorbed water were compressed using an autograph to a thickness of 2 mm. Next, the compressed samples 1 to 4 were depressurized to recover their shape, and the thickness of the recovered samples 1 to 4 was measured. The above procedure was repeated 10 times to measure the shape recovery rate of Samples 1 to 4. The measurement results of the shape recovery rates of Samples 1 to 4 are shown in FIG.

FIG. 6 is a graph showing experimental results of Example 1. The horizontal axis indicates the number of times of compression of each sample, and the vertical axis measures the thickness of the sample after pressure reduction and recovery of the shape after compressing each sample and comparing with the thickness of each sample before compression It shows the shape recovery rate (%) in the case of As shown in FIG. 6, it is confirmed that there is a correlation between the blending ratio (mass ratio) of β-TCP to collagen and the shape recovery rate, and the blending ratio (mass) of β-TCP to collagen It was found that the smaller the ratio, the higher the shape recovery rate. In particular, it was confirmed that in sample 1 (4BT10), sample 2 (9BT10), and sample 3 (13BT10), a high shape recovery rate of about 80% or more can be secured even when the number of compressions increases. On the other hand, it is confirmed that the shape recovery rate of sample 4 (16BT10) after the first time drops to 80% or less, and the shape recovery rate decreases when the compounding ratio (mass ratio) of β-TCP to collagen increases. It was done.

From the above results, by setting the mass ratio of β-TCP to collagen to be 4: 1 or more and less than 10: 1, it is possible to improve the shape recovery of the bone grafting material and tightly close the bone defect. It was also suggested that, even when the bone filling material is filled, it is possible to promote the invasion of blood and osteoblasts into the bone filling material and to promote the new bone formation.

Example 2
(New bone formation ability evaluation)
Next, samples of a total of 10 bone grafting materials 1 in which the blending mass ratio of .beta.-TCP and collagen and the mode diameter of .beta.-TCP particles were changed were prepared, and the new bone forming ability of each prepared samples was compared. .
Specifically, each of the prepared samples A to J was cut out to a diameter of 3 mm and transplanted to a bone defect of 3 mm in diameter formed on the left and right of the rat skull (see FIG. 7). After implantation, the periosteum of the skull was preserved and sutured. Six weeks after implantation, the μCT images were taken to observe the regenerating condition of each sample AJ. The photograph is shown in FIG. FIG. 14 shows SEM images of the cross section of the bone graft material 1 of each of the samples A to J observed at low and high magnifications. Six weeks after transplantation, samples A to J were removed together with the skull surrounding the transplantation site to evaluate bone conductivity and bone penetration. As a comparative example, a case in which the bone defect portion was left as a cavity without implanting the bone filling material 1 was used as a sample K.

The experimental conditions and the results are summarized in Table 1. The comparative experiments were conducted with 3 or 5 samples, and the evaluation was made based on the average value.

New bone regeneration status was mainly confirmed by evaluating bone conductivity and bone penetration. FIGS. 8A to 8C are μCT images showing the determination criteria of each point used to determine bone conductivity, and FIGS. 9A to 9C are μCT showing the determination criteria of each point used to determine bone penetration. It is an image.

As for the evaluation of the bone conductivity, it is confirmed that two points (see FIG. 8A) are partially fused when it is confirmed that the transplanted bone filling material 1 is fused in the entire range. The evaluation was made as 1 point (see FIG. 8B) in the case where it did not occur, and 0 point (see FIG. 8C) when no healing was observed.
For evaluation of bone penetration, two points (see FIG. 9A) are obtained when it is confirmed that the implanted bone graft material 1 is in the center of the bone defect, and the edge of the bone defect It was evaluated as 1 point (see FIG. 9B) when it was confirmed that only new bone was formed, and 0 point (see FIG. 9C) when bone entry was not confirmed. The criteria for bone conductivity and bone penetration are shown in Table 2.

With respect to samples A to K, osteoconductivity and bone infiltration were counted based on the criteria in Table 2 to evaluate the condition of the area in which the new bone was formed in the formed bone defect. The results are shown in FIG.

As a result of this experiment, when the bone defect was left hollow for 6 weeks, the total score of the bone conductivity and the bone penetration was remarkably low at 0.5 point. In the μCT image of sample K after 6 weeks shown in FIG. 10, the bone defect was left as a cavity, and the formation of new bone was hardly confirmed.
In samples C to F and J, where the mode diameter of β-TCP particles of bone filler 1 was 100 to 300 μm, and for sample I, where the mode diameter of β-TCP particles was 50 to 100 μm, β-TCP and The total score of osteoconductivity and bone infiltration was as low as 1 to 2 points regardless of the mass ratio of collagen. In the μCT images of samples C to F, I, and J after 6 weeks shown in FIG. 10, it was confirmed that the healing and bone penetration of the bone graft material 1 filled in the bone defect hardly progressed. .

On the other hand, Sample A in which the mode diameter of the β-TCP particles of the bone grafting material 1 is 1.0 to 2.0 μm and the mass ratio of β-TCP to collagen 1 is 10, β- of the bone grafting material 1 The mode diameter of TCP particles is 1 to 10 μm, the mass ratio of β-TCP to collagen 1 is 10, and the concentration of the crosslinking agent WSC (1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride) is 5.0 wt% In the sample G which is the total score of bone conductivity and bone penetration was as high as 3.7 to 4. In the μCT images of sample A and sample G after 6 weeks shown in FIG. 10 as well, healing of bone grafting material 1 proceeds at the implantation site, and new bone enters the center of the implantation site to regenerate new bone It was confirmed that it was promoted. Sample B in which the mode diameter of the β-TCP particles of the bone grafting material 1 is 1.0 to 2.0 μm and the mass ratio of β-TCP to collagen 1 is 5, the maximum of the β-TCP particles of the bone grafting material 1 Sample H with a frequency of 10-50 μm, a mass ratio of β-TCP to collagen 1 of 10, and a concentration of the crosslinker WSC of 5.0 wt% The In the μCT images of sample B and sample H after 6 weeks shown in FIG. 10, it was confirmed that fusion of bone filling material 1 partially progressed and bone was in the center of the graft site. Thus, it was confirmed that in the sample A, the sample B, the sample G, and the sample H, new bone formation is significantly promoted.

[Example 3]
(Evaluation of component mass ratio)
Next, the component mass ratio of samples A and B of Example 2 was measured. Component mass ratio measurement was performed by calculating the collagen mass which reduced by baking.
Specifically, first, the mass of the crucible used for firing was measured. Next, the mass of sample A (β-TCP / collagen = 10/1 (w / w: at the time of preparation) and sample B (β-TCP / collagen = 5/1 (w / w: at the time of preparation) was measured. Samples A and B were placed in a crucible and fired in a muffle furnace for 10 hours at 1050 ° C. After firing, the weight of β-TCP remaining in samples A and B was measured. Finally, the mass ratio of β-TCP / collagen in sample A and sample B before calcination was calculated.

(Sample A)
<Measurement result>
Mass of crucible A: 152.0460 g
Mass before firing: Mass of crucible A + sample A (β-TCP / collagen composite) = 156.1773 g
Mass after firing: mass of crucible A + sample A (β-TCP) = 155.7560 g
<Calculation result>
Mass of β-TCP + collagen composite: 4.1313 g
β-TCP mass: 3,710 g
Collagen weight: 0.4213 g
β-TCP / collagen mass ratio = 8.81 / 1 (w / w)
(Sample B)
<Measurement result>
Crucible B mass: 10.3905 g
Mass before firing: Mass of crucible B + sample B (β-TCP / collagen composite material) = 10.4572 g
Mass after firing: Mass of crucible B + sample B (β-TCP) = 10.4444 g
<Calculation result>
Mass of β-TCP + collagen composite: 0.0667 g
β-TCP mass: 0.0539 g
Collagen weight: 0.0128 g
β-TCP / collagen mass ratio = 4.21 / 1 (w / w)
From the above, the mass ratio of β-TCP / collagen of sample A and sample B, in which the promoting effect on new bone formation was observed, was 8.81 / 1 (w / w) and 4.21 / 1 (w / respectively). w) confirmed.

Example 4
(Porosity and pore size measurement)
Next, the results of measuring the porosity and pore diameter of the β-TCP particles of the bone grafting material 1 are shown below. The porosity was measured by pore distribution measurement by mercury porosimetry.
First, as a pretreatment, β-TCP particles were vacuum dried at 120 ° C. for 4 hours. Next, the surface tension of mercury was set to 480 dynes / cm, the contact angle between mercury and β-TCP particles was set to 140 degrees, and mercury was made to enter the pores of β-TCP particles. As a measuring device, Autopore IV9520 (micromeritics
Company company) was used.
The measurement results are shown below.
<Measurement result>
Porosity of β-TCP particles: 96%
Pore diameter (median diameter) of β-TCP particles: 141.1 μm
Acceptable range of pore size of β-TCP particles: 20 to 1000 μm
Optimal range of pore size of β-TCP particles: 50 to 500 μm

(Particle size distribution measurement)
Finally, the results of measuring the particle size distribution of β-TCP particles are shown below.
First, as pretreatment, a small amount of β-TCP particles was taken, purified water was added, and the mixture was sonicated for 5 minutes to cancel the aggregation of β-TCP particles. As the measurement conditions, the refractive index of β-TCP was defined as 1.557-0.000i, and the refractive index of water was defined as 1.333. As a measuring device, LS 13320 (BECKMAN COULTER) was used.
The measurement results are shown below.
<Measurement result>
Maximum diameter of β-TCP particles: 15.65 μm

1 Bone filling material 2 Collagen matrix (biodegradable polymer)
3 β-TCP (calcium phosphate)
4 Pore 5 Crosslinking agent SA β-TCP fine powder production process SB Collagen matrix production process SC Bone filling material production process

Claims

The calcium phosphate and the biodegradable polymer are contained in a mass ratio of 4: 1 to 10: 1, and the calcium phosphate includes a plurality of particles having different particle sizes, and the mode diameter of the particles is 20 μm or less.
A porous bone grafting material in which the biodegradable polymer is crosslinked.
The bone grafting material according to claim 1, wherein the pore diameter of the bone grafting material is larger than 20 m.
The bone grafting material according to claim 1 or 2, wherein the porosity of the bone grafting material is 50% or more.
The bone grafting material according to any one of claims 1 to 3, wherein the calcium phosphate is β-tricalcium phosphate (β-TCP).
The bone grafting material according to any one of claims 1 to 4, wherein the biodegradable polymer contains at least collagen.
The bone grafting material according to any one of claims 1 to 5, wherein the crosslinking is a chemical crosslinking.
The bone grafting material according to claim 6, wherein the chemical crosslinking agent used for the chemical crosslinking contains at least one of epoxide and carbodiimide.
Calcining a precursor containing calcium phosphate to form calcium phosphate fine powder;
Mixing the calcium phosphate fine powder with the biodegradable polymer solution and stirring;
A cross-linking step of forming a three-dimensional biodegradable polymer / calcium phosphate composite suspension by adding a cross-linking agent to the product obtained by the stirring step and cross-linking;
Lyophilizing the biodegradable polymer / calcium phosphate composite suspension produced by the crosslinking step;
A washing step of washing away the unreacted crosslinking agent and reaction by-products from the sponge-like biodegradable polymer / calcium phosphate composite porous body obtained by the lyophilization step;
Freezing and drying the washed biodegradable polymer / calcium phosphate composite porous body to produce a bone filling material,
The mass mixing ratio of the calcium phosphate to the biodegradable polymer is 4: 1 to 10: 1,
The method for producing a bone grafting material according to claim 1, wherein the calcium phosphate contains a plurality of particles having different particle sizes, and the mode diameter of the particles is 20 μm or less.
The method for producing a bone graft material according to claim 8, wherein the pore diameter of the bone graft material is larger than 20 μm.
The method for producing a bone graft material according to claim 8 or 9, wherein the porosity of the bone graft material is 50% or more.
The method for producing a bone graft material according to any one of claims 8 to 10, wherein the calcium phosphate is β-tricalcium phosphate (β-TCP).
The method for producing a bone graft material according to any one of claims 8 to 11, wherein the biodegradable polymer contains at least collagen.
The method for producing a bone graft material according to any one of claims 8 to 12, wherein the crosslinking is chemical crosslinking.
The method for producing a bone grafting material according to claim 13, wherein the chemical crosslinking agent used for the chemical crosslinking contains at least one of an epoxide and a carbodiimide.