KR102286084B1 - Control method of curing rate of calcium phosphate Support - Google Patents

Abstract

preparing a paste containing a calcium phosphate-based ceramic; obtaining a molded article by molding the paste prepared above; And the molded body obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ·2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ·2H ₂ O) comprising at least one selected from the group consisting of Disclosed is a method of manufacturing a support for hard tissue regeneration comprising; inducing a cement reaction by immersion in a hardening solution.

Description

Control method of curing rate of calcium phosphate support

The present invention relates to a method for controlling the curing rate of a calcium phosphate support, and more particularly, to a method for manufacturing a support for hard tissue regeneration using a calcium phosphate-based ceramic, and to a support for hard tissue regeneration manufactured through the method.

Tissue engineering, which aims to restore bodily functions lost due to accidents, diseases, and aging, by regeneration rather than replacement treatment, is a multidisciplinary technology that combines various fields such as life science and engineering. is the field Among the three major components of tissue engineering (cells, scaffolds and bioactive molecules), selection of constituent materials and structural control technology are very important for scaffolds. In other words, the scaffold serves as a bridge connecting the tissue to the tissue to regenerate the tissue that has lost its self-repair function. It should have a three-dimensionally well-connected pore structure in a certain size region so that excreta can be exchanged well, and it should have biodegradability that decomposes and disappears according to the rate of tissue regeneration and mechanical strength to maintain its shape during regeneration. and should have excellent biosafety.

In particular, in the regeneration of hard tissues such as bones and teeth, it is important to secure mechanical properties according to the regeneration site. Among the functions required for such a support, the three-dimensional pore structure and mechanical properties are the design and manufacturing technology of the support, and the remaining biodegradability, biocompatibility, and mechanical properties can be controlled mainly by the selection and synthesis of suitable materials.

Representative three-dimensional pore structure manufacturing technologies for scaffolds for hard tissue regeneration include a template method using various polymers or organic materials, a particle leaching method, a gas foaming method using a gas, and fiber meshes. There are methods, phase separation methods, and emulsion freeze drying, but the disadvantages are that it is difficult to control the pore size and connectivity and the reproducibility of the structure is low. 3D printing technology is attracting attention as a process technology to overcome this.

On the other hand, as a material constituting the support, research has been focused on polymers, but in hard tissue regeneration, polymers have a disadvantage in that it is difficult to secure sufficient bioactivity and mechanical properties. Among hard tissues, bone is composed of about 70% inorganic, 20% organic, and 10% water. A study to overcome the shortcomings of polymer materials by using ceramics or ceramic-polymer composites as materials for imitation of the support. Many are in progress. That is, if the ceramic is used as a material and the 3D structure is controlled by 3D printing technology, the development of a support that compensates for the disadvantages of the prior art can be expected.

Representative ceramics used as materials for hard tissue regeneration include calcium phosphate-based ceramics that have a chemical similarity to bone mineral phase. Calcium phosphate ceramics are generally present in powder form, and in order to use this as a raw material to produce a three-dimensional support by 3D printing, a combination of ceramic powder and organic binder is used to form a shape and then high-temperature sintering (>1000°C) to remove organic matter. and forming a bond between the ceramic powder particles to obtain a structurally stable support, or after preparing a three-dimensional shape with a polymer and using it as a template, inject a composite of ceramic powder and an organic binder, and then bake at a high temperature to obtain a polymer and A process for removing the organic binder is applied.

As described above, a high-temperature sintering process is always required to manufacture a three-dimensional support using ceramics. However, heat treatment at high temperature also leads to unstable mechanical properties due to shrinkage and cracking, as well as unexpected crystallization and resulting degradation of biodegradability and bioactivity. Therefore, it is required to develop a low-temperature process without high-temperature heat treatment of the ceramic support. On the other hand, ceramic powder can be bulked by hydration or acid-base reaction through mixing of powder and hardening solution in addition to bulking by high-temperature sintering. Such a reaction is called a cement reaction. Calcium phosphate is a representative bone cement material and is divided into apatite-based and brucite-based materials according to the result. However, in the case of calcium phosphate-based cement, especially brucite-based cement, it is often pointed out that low mechanical strength and strong acidity due to phosphoric acid elution during solution immersion are pointed out as problems. In addition, there is a limit to the structural control of the three-dimensional molded body due to the rapid curing reaction.

As such, a technology for producing calcium phosphate with a controlled three-dimensional shape at a low temperature without conventional high-temperature heat treatment (Korean Patent Publication No. 10-2018-0055960) and a technology for a support for hard tissue regeneration comprising magnesium phosphate (Korea) Patent Registration No. 10-1562556) was developed, but bone cement cannot be directly used for 3D printing due to the rapid curing reaction. Through the induction process, the technical limitations for applying bone cement to 3D printing technology were overcome. However, since a process of immersing the 3D-printed structure in the curing liquid is used without directly mixing the curing liquid with the powder, sufficient time for 3D printing can be secured. If it is a large structure that is too late to penetrate into the structure of the curing liquid, or if the curing reaction is delayed because the contact between the powder and the curing liquid is inhibited due to the introduction of functional substances such as drugs The collapse of the structure may occur, 2) the 3D printed structure may collapse due to the delay of the curing reaction caused by the introduction of functional materials, making it impossible to maintain the shape of the support body, 3) the time required for curing is long, and the process time is long, In addition, 4) there is still a problem related to low mechanical properties that are not sufficient to be used as a support due to insufficient curing reaction.

An object of one aspect of the present invention is to solve the problems associated with the collapse of the 3D printed structure due to the slow curing reaction, long processing time, and insufficient mechanical properties of the support when the bone cement hardening reaction is introduced after 3D printing. there is.

In order to achieve the above object, in one aspect of the present invention

preparing a paste containing a calcium phosphate-based ceramic;

obtaining a molded article by molding the paste prepared above; and

The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Inducing a cement reaction by immersion in a liquid; is provided a method for producing a support for hard tissue regeneration comprising a.

In addition, in another aspect of the present invention

preparing a paste containing a drug including a calcium phosphate-based ceramic and a bisphosphonate-based compound;

obtaining a molded article by molding the paste prepared above; and

Inducing a cement reaction by immersing the molded body obtained above in a curing solution containing MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ·2H _{2 O);}

Furthermore, in another aspect of the present invention

preparing a paste containing a calcium phosphate-based ceramic;

obtaining a molded article by molding the paste prepared above; and

The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Inducing a cement reaction by immersion in a liquid; is provided a support for hard tissue regeneration prepared through a manufacturing method comprising a.

When the method for manufacturing a support for hard tissue regeneration provided in one aspect of the present invention is introduced, it is possible to solve the problem caused by the slow curing reaction of the calcium phosphate-based ceramic. Moreover, it is possible to form a support by resolving the deterioration of the support due to the deterioration of the stability of the support for hard tissue regeneration and delay of the curing reaction caused by the introduction of a functional material, and it is possible to shorten the production time, and the produced support has high mechanical properties.

1 is a flowchart showing a manufacturing method provided in one aspect of the present invention;
2 is a graph showing the hydration reaction-based phase transition time according to the curing solution used in Example 1 and Comparative Example 1;
3 is a graph showing the acid-base reaction-based phase transition time according to the curing solution used in Example 2 and Comparative Example 2;
4 is a graph observing the phase transition according to the curing solution used in Example 1 and Comparative Example 1;
5 is a graph observing the phase transition according to the curing solution used in Example 2 and Comparative Example 2;
6 is a graph showing the reaction completion time according to the curing solution used in Example 1 and Comparative Example 1;
7 is a graph showing the reaction completion time according to the curing solution used in Example 2 and Comparative Example 2;
8 is a photograph of visually observing the supports prepared in Examples 1, 2, Comparative Examples 1 and 2;
9 is a photograph observed by FE-SEM of the supports prepared in Examples 1, 2, Comparative Examples 1 and 2;
10 is a graph summarizing the support curing reaction according to each curing solution in Examples 1, 2, Comparative Examples 1 and 2;
11 is an XRD graph confirming the phase transition according to the curing solution used in Examples 12 to 14 and Comparative Examples 12 to 14;
12 is an XRD graph confirming the phase transition according to the curing solution used in Examples 15 to 17 and Comparative Examples 15 to 17;
13 is an XRD graph confirming the phase transition according to the curing solution used in Examples 18 to 20 and Comparative Examples 18 to 20;
14 is a graph showing the mechanical properties of the supports prepared in Examples 1, 2, Comparative Examples 1 and 2;
15 is a graph of XRD analysis of the support (Example 13 and Comparative Example 13) to which alendronate, a hydrophilic bioactive material, was introduced;
16 is a graph analyzing the cell reactivity of the scaffolds prepared in Examples 1, 2, Comparative Examples 1 and 2;

In one aspect of the invention

preparing a paste containing a calcium phosphate-based ceramic;

obtaining a molded article by molding the paste prepared above; and

The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Inducing a cement reaction by immersion in a liquid; is provided a method for producing a support for hard tissue regeneration comprising a.

At this time, a flowchart showing an example of a method for manufacturing a support for hard tissue regeneration provided in one aspect of the present invention is shown in FIG. 1 ,

Hereinafter, each step will be described in detail with respect to the manufacturing method of the scaffold for hard tissue regeneration provided in one aspect of the present invention.

First, the method for preparing a support for hard tissue regeneration provided in one aspect of the present invention includes preparing a paste containing a calcium phosphate-based ceramic.

In the above step, a paste is prepared using a calcium phosphate-based ceramic material as a ceramic material of the support for hard tissue regeneration.

Here, the calcium phosphate-based ceramic may be a bioceramic raw material capable of inducing a self-curing reaction, α-TCP (α-Tricalcium phosphate), ββTricalcium phosphate), hydroxyapatite (Hydroxyapatite), DCPD (Dicalcium phosphate dihydrate) ), at least one of MCPM (Monocalcium phosphate monohydrate), DCPA (Dicalcium phosphate anhydrous) and BCP (Biphasic Calcium Phosphate) may be used, and preferably α-TCP (α-Tricalcium phosphate) may be used.

The paste is hydroxypropyl methyl cellulose (HPMC), hydroxy ethyl cellulose (HEC), methyl cellulose (Methyl cellulose), carboxymethyl cellulose (CMC), alginate (Alginate), gelatin (Gelatin), collagen (Collagen), five Linogen, Chitosan, Agar, Matrigel, Starch, Pectin, Polyvinyl alcohol, Polyurethane, Polyethylene Gly It may contain at least one thickener of poly (ethylene glycol), polypropylene glycol (Poly (propylene glycol)), hyaluronan (Hyaluronan), and polyvinylpyrrolidone (Poly (vinylpyrrolidone)). there is.

In addition, the content of the thickener may be 0.1 wt% to 5 wt%, 0.5 wt% to 3 wt%, and 0.7 wt% to 2 wt% with respect to the entire paste. By using the thickener in the above range, it is possible to perform molding by imparting appropriate fluidity to the paste, and to secure excellent mechanical properties.

Furthermore, the thickener may include a solvent, and the type of solvent is not particularly limited, but preferably distilled water, tetrahydrofuran, dioxane, ethyl ether, 1,2-dimethoxyethane, dimethylformamide, dimethyl It may further include sulfoxide, dichloromethane, dichloroethane and C ₁ to C ₄ alcohol, and more preferably ethanol. Here, by using the thickener containing the solvent, the viscosity can be adjusted so that the paste can be easily formed and a desired shape is formed. The viscosity of the paste may be preferably adjusted in the range of 10,000 to 1,000,000 cP, preferably in the range of 50,000 to 500,000 cP, and more preferably in the range of 100,000 to 300,000 cP.

Furthermore, the paste may include at least one or more alcohols of methanol, ethanol, propanol, and butanol, and may include an aqueous alcohol solution containing one or more of the alcohols.

In addition, the paste may further include a drug. The drug is Quercetein, Genistein, Curcumin, Saurolactam, Sauchinone, Baicalin, Daidzein, Rutin ), Anthocyanidin, Fisetin, Icarin, Kaempferol, E. Koreanum Nakei, Equol, alendronate, risedronate, etidronate, clodronate, neridronate, ibandronate, zoledronate, olpadronate ) and at least one of a bisphosphonate-based compound. As the paste, quercetin, genestein, etc., which do not affect the phase change occurring during the cement hardening process of ceramics, may be used, but in the present invention, bisphosphonates such as alendronate, including phosphorous, are drugs that affect the cement hardening reaction. (bisphosphonate) may be used.

The content of the drug may be 0.01 wt% to 5 wt%, 0.05 wt% to 3 wt%, and 0.1 wt% to 2 wt% based on the total paste.

The paste may be prepared by pulverizing and mixing calcium phosphate-based ceramics, a drug, and a thickener using a ball mill apparatus. A paste may be prepared by using an aqueous thickener solution further comprising the calcium phosphate-based ceramic and the solvent using a ball mill device. In this case, the average particle size of the powder of the calcium phosphate ceramic may be pulverized to 1 μm to 50 μm, pulverized to 2 μm to 25 μm, and pulverized to 3 μm to 6 μm.

Next, the method of manufacturing a support for hard tissue regeneration provided in one aspect of the present invention includes the step of obtaining a molded body by molding the paste prepared above.

In the above step, the paste prepared in the previous step is molded using various methods to obtain a molded body.

Specifically, the paste prepared above may be obtained as a molded body having a three-dimensional structure through 3D printing technology. The 3D printing technology is a technology for manufacturing a product by continuously reconstructing a digitized three-dimensional product design into a two-dimensional cross section, and then printing raw materials layer by layer.

The thickness of the column of the support can be adjusted by using nozzles of various sizes applied to the 3D printer, and it can be molded into various shapes (pillar spacing, pore size, pore shape, shape of the support, etc.) through a computer program.

Next, the method for producing a support for hard tissue regeneration provided in one aspect of the present invention is the formed body obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO) ₄ ㆍInducing a cement reaction by immersing in a curing solution containing at least one selected from the group consisting of _{2H 2 O).}

Since the molded article is in a state in which the cement reaction has not occurred, it is unstable and easily decomposed in a solution that does not cause a hardening reaction, but it is stable in air and can maintain its shape. Since such a molded body does not use bone cement directly, a hardening reaction is not induced during 3D printing, so sufficient time can be secured for controlling the three-dimensional shape and pore structure, thereby enabling close structural control.

The stabilization of the 3D printed structure is secured by the cement hardening reaction of the raw material powder applied to the paste by immersing it in the curing solution after controlling the three-dimensional shape.

At this time, the conventionally used curing solution has problems such as immediate or sequential collapse of the structure when immersed in the curing solution due to a slow curing reaction, a long process time due to a long curing time, and insufficient mechanical properties to be used as a support. When introducing functional substances such as , drugs, etc., the functional substances delay the reaction between the raw material powder and the curing liquid, so that the 3D printed structure cannot be maintained and there is a problem that it collapses during curing.

Accordingly, in the present invention, by introducing at least one of DSP and MSP as the curing solution, the curing reaction proceeds at a much superior rate than the conventionally used PBS and MCPM (monocalcium phosphates monohydrate), and the problem of structural collapse caused by the introduction of the drug can also be greatly improved.

Specifically, the pH value of the curing liquid may be in the range of 2 to 12. In this case, the pH value of the curing liquid including DSP may be in the range of 8 to 12, and may be in the range of 9 to 11, and the pH value of the curing liquid including MSP may be in the range of 2 to 6, and 3 to It may be in the range of 5. If the pH value of the curing liquid is less than 2, there is a problem of denaturation upon introduction of the functional material due to excessively high acidity, and the curing reaction occurs only on the outside of the 3D structure due to the excessively fast curing reaction, so that a dense layer is formed and the curing liquid reaches the inside The hardening reaction may not be completed due to the difficulty of penetration, and the external layer is peeled off due to the excessive growth of the crystal phase of brusite that is first formed on the outside There is a problem that causes inflammation or cell necrosis.

In addition, the content of at least one of DSP and MSP in the curing solution is preferably 0.1 wt% to 5 wt%, may be 0.5 wt% to 4 wt%, and 1 wt% to 3 wt%, based on the total amount of the curing solution. may be, and may be 2 wt% to 3 wt%. In terms of concentration, the concentration of the curing solution may be 0.01 M to 0.5 M, 0.05 M to 0.4 M, and 0.1 M to 0.2 M. When the concentration range of DSP or MSP in the curing solution is out of the above range, there is a problem in that the phase change is not sufficiently induced or the physical properties of the support cannot be secured due to an excessive phase change.

Furthermore, the immersion of the molded body in the curing solution may be performed for 0.5 hours or more, may be performed for 1 hour or more, may be performed for 3 hours or more, may be performed for 5 hours or more, and may be performed for 6 hours or more. . In addition, it may be performed in 24 hours or less, may be performed in 12 hours or less, and may be performed in 8 hours or less.

As described above, the method for manufacturing a scaffold for hard tissue regeneration provided in one aspect of the present invention can solve the problems caused by the slow curing reaction of calcium phosphate-based ceramics, and reduces the stability of the scaffold for hard tissue regeneration and functional materials It is possible to form a support by resolving the collapse of the support due to the delay of the curing reaction due to the introduction, and to increase the biological functionality of the support, and the produced support has high mechanical properties.

In addition, in another aspect of the present invention

preparing a paste containing a drug including a calcium phosphate-based ceramic and a bisphosphonate-based compound;

obtaining a molded article by molding the paste prepared above; and

Inducing a cement reaction by immersing the molded body obtained above in a curing solution containing MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ·2H _{2 O);}

A bisphosphonate-based compound such as alendronate may interfere with the cement reaction of a molded article formed of a calcium phosphate-based ceramic in the case of a drug containing a phosphorus or phosphate group. In addition, since these hydrophilic drugs contain a large number of OH groups, they may interfere with the curing reaction due to the hydration reaction of the calcium phosphate powder, which causes the 3D structure to collapse without being cured when the curing solution is immersed. In the case of conventional MCPM (monocalcium phosphates monohydrate), an acid-base reaction and a hydration reaction are used at the same time, and compared to a curing reaction using only a hydration reaction of PBS (phosphorus buffer solution) using a simple hydration reaction, the reaction is faster, but still, when introducing a hydrophilic drug The collapse of the structure is confirmed due to insufficient curing reaction. At this time, by applying MSP as the curing solution, the external crystal phase conversion to brusite generated by the rapid curing reaction and acid-base reaction is more stably induced, so that even when a specific drug is included, the stability of the support is reduced or the curing reaction is delayed. It is possible to solve the problem that the structure collapses, and through this, there is an effect of improving mechanical properties. In the case of using only a hydration reaction such as conventional PBS, the above-mentioned problems can be solved by inducing a fast hydration reaction by converting the solution to DSP.

Furthermore, in another aspect of the present invention

preparing a paste containing a calcium phosphate-based ceramic;

obtaining a molded article by molding the paste prepared above; and

The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Inducing a cement reaction by immersion in a liquid; is provided a support for hard tissue regeneration prepared through a manufacturing method comprising a.

The support for hard tissue regeneration provided in another aspect of the present invention is a support for hard tissue regeneration prepared by solving problems caused by the slow curing reaction of calcium phosphate-based ceramics, and exhibits high mechanical properties.

Hereinafter, the present invention will be described in detail through Examples and Experimental Examples.

However, the following examples and experimental examples are only for illustrating the present invention, and the content of the present invention is not limited by the following examples and experimental examples.

<Example 1> Preparation of a support using DSP as a curing liquid 1

Step 1: α-TCP (α-Tricalcium phosphate) powder is mixed with a 30% aqueous ethanol solution containing 1 wt% of hydroxypropyl methyl cellulose by adjusting the separation ratio of 1:0.5 - 1:0.67, and representative The paste was prepared using a 3D printer ME (material extruding) system.

Step 2: Put the paste prepared in Step 1 into an extrusion container, and use a 3D printer to prepare a three-dimensional support, and then dry it at a temperature of 37° C. for 24 hours.

Step 3: Using an aqueous solution of 0.14 M DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) as a curing solution, the support prepared in step 2 was immersed in the solution for 0 to 48 hours to induce a curing reaction. .

Step 4: After washing the support prepared in step 3 with distilled water several times, it was dried to prepare a support.

<Example 2> Preparation of a support using MSP as a curing solution 2

A support was prepared in the same manner as in Example 1, except that in step 3 of Example 1, 0.16 M MSP (Disodium phosphate dihydrate, Na ₂ HPO ₄ ·2H _{2 O) aqueous solution was used as the curing solution.}

<Example 3-5> Preparation of quercetin-containing support using DSP as a curing solution 1-3

A support was prepared in the same manner as in Example 1, except that Quercetein was mixed with the paste in Step 1 of Example 1, wherein the content of quercetin was 0.1 wt%, 0.5 wt% and 1.0 wt%, respectively %am.

< Example 6-8> with hardening liquid using DSP Genestein Preparation of the containing support 1-3

A support was prepared in the same manner as in Example 1, except that Genistein was mixed with the paste in Step 1 of Example 1, wherein the content of genestein was 0.1 wt%, 0.5 wt%, and 1.0% by weight.

< Example 9-11> with hardening liquid Preparation of alendronate-containing support using DSP 1-3

A support was prepared in the same manner as in Example 1, except that alendronate was mixed with the paste in step 1 of Example 1, wherein the content of alendronate was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively. %am.

<Example 12-14> Preparation of a support containing quercetin using MSP as a curing solution 1-3

A support was prepared in the same manner as in Example 2, except that Quercetein was mixed with the paste in Step 1 of Example 2, wherein the content of quercetin was 0.1% by weight, 0.5% by weight, and 1.0% by weight, respectively. %am.

< Example 15-17> with hardening liquid msp used Genestein Preparation of the containing support 1-3

A support was prepared in the same manner as in Example 2, except that Genistein was mixed with the paste in Step 1 of Example 2, wherein the contents of Genestein were 0.1 wt%, 0.5 wt%, and 1.0% by weight.

< Example 18-20> with hardening liquid msp Preparation of the used alendronate-containing support 1-3

A support was prepared in the same manner as in Example 2, except that alendronate was mixed with the paste in step 1 of Example 2, wherein the content of alendronate was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively. %am.

<Comparative Example 1> Preparation of a support using PBS as a curing solution

A support was prepared in the same manner as in Example 1, except that in step 3 of Example 1, 1X aqueous PBS (Phosphate buffer saline) was used as the curing solution.

<Comparative Example 2> Preparation of a support using MCPM as a curing liquid

A support was prepared in the same manner as in Example 1, except that 0.1 M aqueous solution of monocalcium phosphates monohydrate (MCPM) was used as the curing solution in step 3 of Example 1.

<Comparative Example 3-5> Preparation of a support containing quercetin using PBS as a curing solution 1-3

A support was prepared in the same manner as in Comparative Example 1 except that Quercetein was mixed with the paste in Step 1 of Comparative Example 1, wherein the content of quercetin was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively %am.

<Comparative Example 6-8> Preparation of a support containing genesteine using PBS as a curing solution 1-3

A support was prepared in the same manner as in Comparative Example 1, except that in step 1 of Comparative Example 1, genestein was mixed with the paste, wherein the content of genestein was 0.1 wt%, 0.5 wt%, and 1.0% by weight.

<Comparative Example 9-11> Preparation of an alendronate-containing support using PBS as a curing solution 1-3

A support was prepared in the same manner as in Comparative Example 1, except that alendronate was mixed with the paste in Step 1 of Comparative Example 1, wherein the content of alendronate was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively. %am.

<Comparative Example 12-14> Preparation of a support containing quercetin using MCPM as a curing solution 1-3

A support was prepared in the same manner as in Comparative Example 2, except that Quercetein was mixed with the paste in Step 1 of Comparative Example 2, wherein the content of quercetin was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively %am.

<Comparative Example 15-17> Preparation of a support containing genesteine using MCPM as a curing solution 1-3

A support was prepared in the same manner as in Comparative Example 2, except that Genistein was mixed with the paste in Step 1 of Comparative Example 2, wherein the content of Genestein was 0.1 wt%, 0.5 wt%, and 1.0% by weight.

<Comparative Example 18-20> Preparation of an alendronate-containing support using MCPM as a curing liquid 1-3

A support was prepared in the same manner as in Comparative Example 2, except that alendronate was mixed with the paste in step 1 of Comparative Example 2, wherein the content of alendronate was 0.1 wt%, 0.5 wt%, and 1.0 wt%, respectively. %am.

<Experimental example>

As shown in FIG. 2, looking at the hydration reaction-based phase transition to calcium deficient hydroxyapatite (CDHA), compared to Comparative Example 1 using PBS as the curing solution, Example 1 using DSP as the curing solution was faster. It can be confirmed that the curing reaction is shown. In the case of PBS, it was confirmed that at least 12 hours were required for complete phase transition to CDHA, whereas in the case of DSP, complete phase transition was possible in 6 hours. That is, the curing process time can be reduced in half.

As shown in Figure 3, looking at the acid-base reaction and the hydration reaction-based dicalcium phosphate dihydrate (DCPD)-hydroxyapatite (calcium deficient hydroxyapatite, CDHA) phase transition, the implementation using MSP as a curing solution In Example 2 and Comparative Example 2 using MCPM as the curing solution, DCPD was generated at a similar rate, but in the case of CDHA, it was confirmed that MSP was generated faster than MCPM. That is, faster internal structure hardening and structural stability can be secured through this.

As shown in Figures 4 and 6, it was possible to observe a phenomenon in which the reaction was terminated faster in DSP than in PBS.

As shown in FIGS. 5 and 7 , it was observed that the reaction was terminated faster in MSP than in MCPM.

8 is a photograph observed with the naked eye of the supports prepared in Examples 1, 2, Comparative Examples 1 and 2, and FIG. 9 is prepared in Examples 1, 2, Comparative Example 1 and Comparative Example 2 It is a photograph of the observed support with FE-SEM.

In addition to FIGS. 8 and 9, as shown in the graph of FIG. 10 summarizing the XRD, FE-SEM and ion elution results, when DSP (Example 1) is used than when using PBS (Comparative Example 1) It could be confirmed that curing was rapidly induced, and it was confirmed that when using MSP (Example 2), the support curing reaction was induced faster than when using MCPM (Comparative Example 2).

11 to 13 show the results of observing the curing reaction according to the hydrophilicity/hydrophobicity of the drug for the support on which the three kinds of drugs were supported. 11 to 13, Q means a support on which the hydrophobic drug quercetin is supported, G means a support on which the hydrophobic drug Genestein is supported, A is a hydrophilic drug and a support on which alendronate including a phosphorous group is supported. it means.

As shown in FIG. 11, since quercetin is a hydrophobic material and is a drug that does not contain factors affecting the cement hardening reaction, it is confirmed that the support hardening reaction is induced regardless of the drug loading concentration, so that a-TCP is replaced with CDHA. could As shown in FIG. 12 , even in the case of Genestein, since it is a hydrophobic substance and is a drug that does not contain factors affecting the cement hardening reaction, the cement hardening reaction in which a-TCP is replaced with CDHA proceeds regardless of the drug loading concentration. It was confirmed that the support hardening reaction was induced. On the other hand, as shown in FIG. 13, in the case of alendronate, it contains a phosphorus group that affects the cement hardening reaction and is a hydrophilic material, so it interferes with the hydration reaction used for hardening the support. Although the curing reaction occurs regardless of the type, it is confirmed that the CDHA phase is generated the fastest in MSP, and in the case of 0.5% by weight of the drug, which is 5 times higher, the curing reaction does not occur in the conventional reaction in PBS, so only the a-TCP phase This is observed, and in the case of MCPM, it is confirmed that the curing reaction is not completed and some a-TCP is substituted only with DCPD due to an acid-base reaction that occurs preferentially, and not with CDHA, which is the result of the hydration reaction. On the other hand, when the MSP presented in the present invention was used as the curing solution, it was confirmed that all a-TCPs were substituted with CDHA to induce a complete curing reaction.

As shown in FIG. 14, it was confirmed that the mechanical properties of the support reacted in DSP were improved compared to the support reacted in PBS, and the mechanical properties of the support reacted in MSP were improved compared to the support reacted in MCPM. It was confirmed that the structure exhibited the highest mechanical properties.

That is, it was confirmed that mechanical properties could be improved by controlling the curing reaction rate.

In the case of a hydration reaction (transition to CDHA), the reaction may be inhibited or prevented by introduction of a hydrophilic bioactive material. That is, the 3D printed structure cannot be maintained because it cannot be cured due to failure to induce the cement reaction of the 3D printed structure.

When alendronate, a hydrophilic material, is directly mixed with a-TCP, in the case of MCPM and MSP, which induce an acid-base reaction and a hydration reaction simultaneously, a curing reaction is induced and a structure is induced, but a-TCP, which has not been transferred after 24 hours of reaction, is In contrast to the remaining MCPM, in the case of MSP, which induces a fast curing reaction, a slow curing reaction is induced compared to before the introduction of alendronate, but it can be confirmed that a stable structure can be formed by completely transferring to CDHA and DCPD (see Fig. 15).

That is, it was confirmed that the support could be stably manufactured without collapse of the 3D structure due to the introduction of the bioactive material by controlling the curing reaction rate.

Furthermore, it was confirmed that only by the manufacturing method of the present invention, which rapidly controls the cement hardening rate during hardening of the bone scaffold, alendronate (bisphosphonate-based drug), which is a hydrophilic material and a representative treatment for osteoporosis, can be stably supported.

As shown in FIG. 16 , there was no significant difference in proliferation in the case of scaffolds cured in a basic DSP reaction solution compared to PBS having a neutral pH, but it was confirmed that differentiation was somewhat lower. However, compared to MCPM, which is a reaction solution for a strong acid region, MSP, a weak acid reaction solution, showed excellent cell proliferation and differentiation behavior.

As a result of the above results, it is most preferable to use MSP, which can induce a fast curing rate in terms of stability to the support, mechanical properties, ease of introduction of bioactive materials, and cell reactivity, as the curing solution, and DSP and MSP are mixed with PBS and MCPM solution. It has been verified that the problem of the existing reaction mentioned above can be solved by using it instead of .

Claims (13)

preparing a paste containing a calcium phosphate-based ceramic;
obtaining a molded article by molding the paste prepared above; and
The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Including; inducing a cement reaction by immersing in the liquid
The concentration of one or more selected from the group consisting of disodium phosphate dihydrate, Na ₂ HPO ₄ .2H ₂ O) and monosodium phosphate dihydrate (MSP, NaH ₂ PO ₄ .2H _{2 O) in the curing solution is 0.01 M to 0.5} Method for producing a scaffold for hard tissue regeneration, characterized in that M.
According to claim 1,
The paste is a method of manufacturing a scaffold for hard tissue regeneration further comprising a drug.
According to claim 1,
The calcium phosphate-based ceramic is α-TCP (α-Tricalcium phosphate), β-TCP (β-Tricalcium phosphate), hydroxyapatite (Hydroxyapatite), DCPD (Dicalcium phosphate dihydrate), MCPM (Monocalcium phosphate monohydrate), DCPA (Dicalcium) Phosphate anhydrous) and BCP (Biphasic Calcium Phosphate) method for producing a scaffold for hard tissue regeneration comprising at least one selected from the group consisting of.
3. The method of claim 2,
The drug is a method for producing a scaffold for hard tissue regeneration comprising at least one selected from the group consisting of bisphosphonate compounds.
According to claim 1,
The paste is hydroxypropyl methyl cellulose (HPMC), hydroxy ethyl cellulose (HEC), methyl cellulose (Methyl cellulose), carboxymethyl cellulose (CMC), alginate (Alginate), gelatin (Gelatin), collagen (Collagen), five Linogen, Chitosan, Agar, Matrigel, Starch, Pectin, Polyvinyl alcohol, Polyurethane, Polyethylene Gly Col (Poly (ethylene glycol)), polypropylene glycol (Poly (propylene glycol)), hyaluronan (Hyaluronan) and polyvinylpyrrolidone (Poly (vinylpyrrolidone)) at least one selected from the group consisting of Method for producing a scaffold for hard tissue regeneration further comprising an agent.
According to claim 1,
The paste is a method of manufacturing a support for hard tissue regeneration further comprising at least one alcohol selected from the group consisting of methanol, ethanol, propanol and butanol.
3. The method of claim 2,
The content of the drug is 0.01% by weight to 5% by weight of the total paste manufacturing method of the scaffold for hard tissue regeneration.
According to claim 1,
The molding is a method of manufacturing a support for hard tissue regeneration that is performed by a layered molding method.
According to claim 1,
The pH value of the hardening solution is in the range of 2 to 14. Method for producing a support for hard tissue regeneration.
delete According to claim 1,
The method for producing a support for hard tissue regeneration is performed for 0.5 to 8 hours by immersing the molded body in the hardening solution.
preparing a paste containing a drug including a calcium phosphate-based ceramic and a bisphosphonate-based compound;
obtaining a molded article by molding the paste prepared above; and
Inducing a cement reaction by immersing the molded body obtained above in a curing solution containing MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H _{2 O);}
The concentration of one or more selected from the group consisting of disodium phosphate dihydrate, Na ₂ HPO ₄ .2H ₂ O) and monosodium phosphate dihydrate (MSP, NaH ₂ PO ₄ .2H _{2 O) in the curing solution is 0.01 M to 0.5} Method for producing a scaffold for hard tissue regeneration, characterized in that M.
preparing a paste containing a calcium phosphate-based ceramic;
obtaining a molded article by molding the paste prepared above; and
The molded article obtained above, DSP (disodium phosphate dihydrate, Na ₂ HPO ₄ ㆍ2H ₂ O) and MSP (monosodium phosphate dihydrate, NaH ₂ PO ₄ ㆍ2H ₂ O) curing comprising at least one selected from the group consisting of Including; inducing a cement reaction by immersing in the liquid
The concentration of one or more selected from the group consisting of disodium phosphate dihydrate, Na ₂ HPO ₄ .2H ₂ O) and monosodium phosphate dihydrate (MSP, NaH ₂ PO ₄ .2H _{2 O) in the curing solution is 0.01 M to 0.5} A support for hard tissue regeneration manufactured through a manufacturing method, characterized in that M.

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