阅读说明：本技术 骨移植组合物及其制备方法 (Bone graft composition and method for preparing the same ) 是由 朴庭馥 于 2020-06-24 设计创作，主要内容包括：本公开涉及一种包含羟丙基甲基纤维素的骨移植组合物及其制备方法。更具体地,一种骨移植组合物及其制备方法,所述骨移植组合物包含提供最佳渗透性质和形状保持性的量的羟丙基甲基纤维素。(The present disclosure relates to a bone graft composition comprising hydroxypropylmethylcellulose and a method for preparing the same. More particularly, a bone graft composition comprising hydroxypropylmethylcellulose in an amount to provide optimal osmotic properties and shape retention, and a method of making the same.)

1. A bone graft composition, wherein, when the osmotic pressure of saline is set to 100% as a reference value, the saline reaches 104 to 112% of the osmotic pressure within 12 to 48 hours after the addition of the bone graft solution.

2. The bone graft composition according to claim 1, wherein the bone graft solution is a mixture of 1 part by weight of a bone graft material comprising hydroxypropylmethylcellulose and 0.5 to 2 parts by weight of a solvent, and the bone graft material comprising hydroxypropylmethylcellulose is formed by mixing 1 part by weight of the bone graft material with 0.3 to 3 parts by weight of hydroxypropylmethylcellulose.

3. The bone graft composition of claim 2, wherein the bone graft material is a natural bone graft material.

4. The bone graft composition according to claim 2, wherein the solvent is water.

5. A method for preparing a bone graft composition, the method comprising the steps of:

(1) preparing a bone morphogenetic protein solution by mixing a solvent and the bone morphogenetic protein;

(2) adsorbing the bone morphogenetic protein onto the graft material powder by mixing the bone morphogenetic protein solution with the graft material powder;

(3) mixing and stirring a graft material powder having the bone morphogenetic protein adsorbed thereon and a hydroxypropyl methylcellulose powder to form a gel having osmotic properties; and

(4) the structure comprising the plurality of wells is formed by freeze-drying the gel under vacuum.

6. The method of claim 5, wherein the bone morphogenic protein is at least one selected from the group consisting of BMP-2, BMP-3b, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, BMP-9, BMP-10, BMP-11, BMP-12, BMP-13, BMP-14, BMP-15, BMP-16, BMP-17, BMP-18, recombinant bone morphogenic proteins thereof, and equivalents thereof.

7. The method of claim 5, wherein the concentration of the bone morphogenic protein in the bone morphogenic protein solution is 0.05mg/mL to 0.15 mg/mL.

8. The method of claim 5, wherein the pH of the bone morphogenetic protein solution is adjusted to 4.6 to 5 using phosphate buffered saline.

9. The method according to claim 5, wherein the volume ratio between the graft material powder having the bone morphogenetic protein adsorbed thereon in the step (3) and the hydroxypropylmethylcellulose powder is 1:0.2 to 1: 0.6.

10. The method of claim 5, further comprising the step of sterilizing the bone graft composition by ethylene oxide gas or gamma irradiation.

11. The method according to claim 10, wherein the concentration of the ethylene oxide gas is 450mg/L to 1,200mg/L, or the dose of the gamma ray irradiation is 10kGy to 25 kGy.

Technical Field

The present disclosure relates to a bone graft composition and a method of preparing the same.

Background

Various materials and methods may be used to reconstruct defective bone. For example, for reconstruction of a defective bone, a bone graft material such as bone powder, bone fragments, and bone blocks may be used, or a method such as autograft, allograft, and xenograft may be used.

Bone graft materials for the reconstruction of defective bones can be used in bone surgery, neurosurgery, orthopedics, otorhinolaryngology, oral and maxillofacial surgery, veterinary (veterinary clinics), dermatology and dentistry. For example, these materials may be used for bone defects during disc surgery to induce bone regeneration, or may also be used for implantation surgery and reconstruction of oral and maxillofacial bone defects.

In addition, Korean patent No. 10-0401941 discloses a technology related to a bone graft material and a method for preparing the same. As disclosed therein, when a mesh bone composed of a bioceramic powder and having a three-dimensional interconnected pore structure is used, the effect of bone grafting may be limited in terms of biocompatibility, mechanical properties, toxicity, and the like.

Disclosure of Invention

The present disclosure is directed to providing a bone graft composition suitable for bone formation, including hydroxypropylmethylcellulose, and particularly, a bone graft composition and a bone graft solution that form an optimal osmotic pressure with another solution.

One embodiment of the present disclosure provides a bone graft composition, wherein, when the osmotic pressure of saline is set to 100% as a reference value, the saline reaches 104 to 112% of the osmotic pressure within 12 to 48 hours after the addition of a bone graft solution.

One embodiment of the present disclosure provides a bone graft composition, wherein the bone graft solution is a mixture of 1 part by weight of a bone graft material including hydroxypropylmethylcellulose and 0.5 to 2 parts by weight of a solvent, and the bone graft material including the hydroxypropylmethylcellulose is formed by mixing 1 part by weight of the bone graft material with 0.3 to 3 parts by weight of the hydroxypropylmethylcellulose.

One embodiment of the present disclosure provides a bone graft composition, wherein the bone graft material is a natural bone graft material.

One embodiment of the present disclosure provides a bone graft composition, wherein the solvent is water.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, the method comprising the steps of: (1) preparing a bone morphogenetic protein solution by mixing a solvent and the bone morphogenetic protein; (2) adsorbing the bone morphogenetic protein onto the graft material powder by mixing the bone morphogenetic protein solution with the graft material powder; (3) mixing and stirring a graft material powder having the bone morphogenetic protein adsorbed thereon and a hydroxypropyl methylcellulose powder to form a gel having osmotic properties; and (4) forming a structure comprising a plurality of pores by freeze-drying the gel under vacuum.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the bone morphogenetic protein is at least one selected from the group consisting of BMP-2, BMP-3b, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, BMP-9, BMP-10, BMP-11, BMP-12, BMP-13, BMP-14, BMP-15, BMP-16, BMP-17, BMP-18, recombinant bone morphogenetic proteins thereof, and bone morphogenetic proteins equivalent thereto.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the concentration of bone morphogenic protein in the bone morphogenic protein solution is 0.05mg/mL to 0.15 mg/mL.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the pH of a bone morphogenic protein solution is adjusted to 4.6 to 5 using phosphate buffered saline.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the volume ratio between the graft material powder having bone morphogenetic protein adsorbed thereon and the hydroxypropylmethylcellulose powder in step (3) is 1:0.2 to 1: 0.6.

An embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the method further comprises the step of sterilizing the bone graft composition by ethylene oxide gas or gamma ray irradiation.

One embodiment of the present disclosure provides a method for preparing a bone graft composition, wherein the concentration of ethylene oxide gas is 450mg/L to 1,200mg/L, or the dose of gamma ray irradiation is 10kGy to 25 kGy.

One embodiment of the present disclosure provides a bone graft composition prepared by the above-described method for preparing a bone graft composition.

Drawings

Fig. 1 is a flow diagram schematically illustrating a method for preparing a bone graft composition according to one embodiment of the present disclosure.

Fig. 2 shows result data of an experimental example of the present disclosure, which is obtained by mixing saline with a bone graft solution formed using different amounts (parts by weight) of a solvent and expressing a change in osmotic pressure of the mixed saline as compared to that of pure saline over time as an osmotic rate.

Detailed Description

Embodiments of the present disclosure relate to a bone graft composition that may have excellent effects in activating bone formation, biocompatibility, and ease of use by including a porous bone graft material and hydroxypropylmethylcellulose.

However, descriptions of parts of the specific embodiments that overlap with the descriptions of the other embodiments will be omitted for a clearer and more concise description. Even if the description of the portion is omitted, the portion is not excluded from the present disclosure, and the scope of the right thereof should be recognized in the same manner as the other embodiments.

In the following description, a detailed description of well-known technologies related to the present disclosure will be omitted when it may unnecessarily obscure the subject matter of the present disclosure. In addition, terms used in the following description are terms defined in consideration of their functions in the present disclosure, and may be changed according to the intention of a user or an operator or according to practice. Therefore, the definitions of these terms should be determined based on the contents throughout the specification.

The technical spirit of the present disclosure is determined by the claims, and the following examples are only means for effectively explaining the technical spirit of the present disclosure to those skilled in the art to which the present disclosure pertains.

In the present disclosure, when the repeating unit, compound or resin represented by the formula includes isomers thereof, the corresponding formula representing the repeating unit, compound or resin means a representative formula also representing isomers.

Hereinafter, specific embodiments of the present disclosure will be described. However, these embodiments are merely examples, and the present disclosure is not limited thereto.

The bone graft composition may be implanted into a bone defect and may be used to repair a bone defect by filling the bone defect. Hereinafter, "implantation" includes administration into a bone defect in a state without rigidity or in a state with rigidity. The shape-forming device (e.g., a three-dimensional printer) may be used to form a shape corresponding to the shape of the bone defect.

The bone graft composition of the present disclosure comprises a bone graft material and hydroxypropyl methylcellulose. The bone graft composition may be implanted into a bone defect and may be used to repair a bone defect by filling the bone defect.

The bone graft material may be natural bone, such as autologous bone, allogeneic bone or xenogeneic bone. When natural bone is used, the natural bone may exhibit an excellent bone forming effect because the natural bone has excellent biocompatibility and also has good wettability and hygroscopicity due to a large number of pores contained therein. In addition, natural bone can also be used for reconstruction of defective bone in bone surgery, neurosurgery, orthopaedics, otorhinolaryngology, oral and maxillofacial surgery, veterinary medicine (veterinary clinic), dermatology and dentistry.

In addition, bone graft materials may also be used for the reconstruction of defective bones in humans or animals. Hereinafter, the use for dentistry is mainly described, however, the use is not limited thereto.

Since the bone graft composition includes hydroxypropylmethylcellulose, the bone graft composition may have adhesiveness to the bone defect. When the bone graft composition has excellent adhesiveness, it does not flow downward even if the bone graft composition is applied to the maxilla, and the bone graft composition can be prevented from falling off from a bone defect even if there is an impact due to a chewing motion.

In addition, since the bone graft composition includes hydroxypropylmethylcellulose, hydroxypropylmethylcellulose may affect the formation of osmotic pressure between the composition and another solution. Depending on the degree of dissolution of the hydroxypropylmethylcellulose in the solvent, a change in osmotic pressure occurs between the composition and another solution. When the bone graft composition is applied to a bone defect under conditions that create an optimal osmotic pressure, a medical procedure for applying the composition can be more easily performed by minimizing the influence of external conditions.

According to one embodiment of the present disclosure, a bone graft composition exhibiting optimal bone graft properties during a medical procedure may include hydroxypropylmethylcellulose in an amount of 0.15 to 6 parts by weight (more preferably, 0.3 to 3 parts by weight) based on 1 part by weight of a bone graft material. If the content of the hydroxypropylmethylcellulose is less than 0.3 parts by weight based on 1 part by weight of the bone graft material, the composition has insufficient adhesion to the bone defect and thus is highly likely to fall off from the bone defect during use, and since the content of the hydroxypropylmethylcellulose is insignificant, the hydroxypropylmethylcellulose has little or no effect on the osmotic property of the bone graft composition. If the content of the hydroxypropylmethylcellulose is more than 3 parts by weight per part by weight of the bone graft material, the hydroxypropylmethylcellulose may interfere with bone formation by interfering with wettability, hygroscopicity, and the like of alien bone, and the hydroxypropylmethylcellulose may not be easily dissolved due to solidification thereof, and may have a great influence on osmotic pressure of the composition due to outward flow thereof, so that the composition may not be suitable for use as a bone graft material.

The bone graft composition comprising hydroxypropylmethylcellulose is used as a solution obtained by dissolving it in a solvent. Any solvent that can be used in the art may be appropriately selected and used as the solvent, for example, water may be used as the solvent. Since the physical properties of the bone graft composition, such as dissolution rate, concentration, osmotic properties, and shape retention, vary depending on the conditions of the solvent, it is important to select appropriate solvent conditions for the practice of the present disclosure.

According to one embodiment of the present disclosure, in order for a bone graft solution to exhibit optimal properties during a medical procedure (in particular, in order for a bone graft solution to form an optimal osmotic pressure with another solution), a bone graft solution may be prepared by mixing 1 part by weight of a bone graft composition including hydroxypropylmethylcellulose with 0.5 to 2 parts by weight of a solvent (water). As can be seen from experimental examples to be described later, the content of the solvent should be in a range such that the bone graft solution can form a proper permeation phenomenon with another solution. As shown in fig. 2, it can be seen that 0.5 to 2 parts by weight of a solvent (water) should be mixed with 1 part by weight of a bone graft composition comprising hydroxypropylmethylcellulose in order to form a proper permeation phenomenon.

If the content of the solvent (water) is less than 0.5 parts by weight based on 1 part by weight of the bone graft composition comprising hydroxypropylmethylcellulose, since the amount of the solvent (water) is too small, dissolution may not easily occur, and thus hydroxypropylmethylcellulose may not be easily combined with the bone graft material and may not sufficiently exhibit its function. Generally, the composition may be too hard for it to be suitable for bone formation. In addition, it may form a strong osmotic pressure with another solution, so that the bone graft material does not maintain its shape and its function may be impaired.

If the content of the solvent (water) is more than 2 parts by weight based on 1 part by weight of the bone graft composition including hydroxypropylmethylcellulose, the amount of the solvent (water) is too large, so that the solution has a flowing property without having a suitable viscosity, and thus the composition may not have shape-retaining properties. In addition, the dissolution of hydroxypropylmethylcellulose proceeds rapidly due to a high content of solvent (water), and thus hydroxypropylmethylcellulose may flow outward, so that it cannot exhibit its function. In addition, it may form a strong osmotic pressure with another solution, so that the bone graft material does not maintain its shape and its function may be impaired.

Therefore, as solvent conditions for allowing the bone graft composition comprising hydroxypropylmethylcellulose to sufficiently exhibit the function of hydroxypropylmethylcellulose and maintain the shape of the bone graft material, 0.5 to 2 parts by weight of a solvent (water) was mixed with 1 part by weight of the bone graft composition comprising hydroxypropylmethylcellulose to prepare a bone graft solution.

A bone graft composition kit according to another embodiment of the present disclosure includes the above bone graft composition and a syringe containing the composition. By providing a syringe that directly contains the bone graft composition, ease of use can be assured and the likelihood of contamination that may occur during use is significantly reduced.

However, in the description of the present embodiment, descriptions of parts overlapping with those of the other embodiments are omitted for clearer and more concise description. Even if the description of the portion is omitted, the portion is not excluded from the present disclosure, and the scope of the right thereof should be recognized in the same manner as the other embodiments.

A method for preparing a bone graft composition according to yet another embodiment of the present disclosure includes the steps of:

1. dissolving a bone morphogenetic protein to prepare a bone morphogenetic protein solution;

2. dropping the graft material powder into the bone morphogenetic protein solution prepared by the step 1 to impregnate; or

3. Stirring after adding the bone morphogenetic protein solution prepared in the step 1 to the graft material powder to allow the bone morphogenetic protein to be adsorbed;

4. mixing and stirring the bone morphogenetic protein solution and graft material powder prepared in step 2 or step 3 with hydroxypropylmethylcellulose (powder form) to form a gel shape; and

5. the sponge-like structure comprising a plurality of pores was formed by freeze-drying a mixture of graft material powder and hydroxypropyl methylcellulose powder obtained by a mixing and stirring process at a low temperature under vacuum. The bone graft composition prepared through these steps may have excellent effects in activating bone formation, biocompatibility, and ease of use.

However, in the description of this embodiment, the description of the parts overlapping with those of the above-described embodiment is omitted for the sake of clearer and more concise explanation. Even if the description of the portion is omitted, the portion is not excluded from the present disclosure, and the scope of the right thereof should be recognized in the same manner as the above-described embodiments.

Fig. 1 is a flow diagram schematically illustrating a method for preparing a bone graft composition according to one embodiment of the present disclosure.

Fig. 2 shows the result data of the experimental example of the present disclosure, which is obtained by mixing saline with a bone graft solution formed using different amounts (parts by weight) of a solvent, and then expressing the change in osmotic pressure of the mixed saline with time as compared with that of pure saline as permeability.

First, a bone morphogenic protein solution is prepared by dissolving the bone morphogenic protein in a solvent. The bone morphogenic protein solution can be prepared by adding the bone morphogenic protein to the solvent or adding the bone morphogenic protein to the solvent and dissolving the bone morphogenic protein in the solvent.

The bone morphogenic protein can be at least one selected from the group consisting of BMP-2, BMP-3b, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, BMP-9, BMP-10, BMP-11, BMP-12, BMP-13, BMP-14, BMP-15, BMP-16, BMP-17, BMP-18, recombinant bone morphogenic proteins thereof, and equivalent bone morphogenic proteins thereof. For the bone forming effects of the present disclosure, preferably, the bone morphogenetic protein can be rhBMP-2.

According to one embodiment of the present disclosure, the concentration of the bone morphogenetic protein in the bone morphogenetic protein solution can be 0.05mg/mL to 0.15mg/mL, preferably, can be 0.08mg/mL to 0.12 mg/mL. When the concentration of the bone morphogenetic protein is within the above range, bone formation by the bone morphogenetic protein can be activated. If the concentration of the bone morphogenetic protein is less than 0.05mg/mL, the ability of the bone morphogenetic protein to form new bone is reduced, and if the concentration of the bone morphogenetic protein is more than 0.15mg/mL, it causes adverse effects.

In addition, according to one embodiment of the present disclosure, the pH of the bone morphogenic protein solution can be, for example, 4.6 to 5. When the pH is within the above range, bone formation via bone morphogenetic proteins can be activated. If the pH of the bone morphogenetic protein solution is less than 4.6, the ability to form new bone is reduced, and if the pH of the bone morphogenetic protein solution is greater than 5, the ability to form new bone is reduced. For example, phosphate buffered saline may be used to adjust the pH. When pH is adjusted using phosphate buffered saline, the bone morphogenic protein can have the effect of forming new bone.

Thereafter, bone morphogenic protein is adsorbed onto the graft material powder by soaking the graft material powder with a bone morphogenic protein solution. The bone morphogenic protein can be adsorbed onto the graft material powder by soaking the graft material powder with the bone morphogenic protein solution by rinsing the previously prepared graft material powder with the bone morphogenic protein solution or dropping the graft material powder into the bone morphogenic protein solution.

The graft material powder may be autologous bone, allogeneic bone or xenogeneic bone. For example, the graft material powder may be prepared by placing the graft material powder in a snap (or so-called snap or snap) tube.

The average particle diameter (D50) of the graft material powder may be 200 μm to 5,000 μm, and preferably, may be 250 μm to 1,000 μm. If the mean particle diameter of the graft material powder is less than 200 μm, the graft material may be rapidly absorbed, so bone conduction required for bone formation may be insufficient, and if the mean particle diameter of the graft material powder is greater than 5,000 μm, precise handling of the graft material powder during administration to a patient may be difficult.

According to one embodiment of the present disclosure, the step of adsorbing bone morphogenic protein onto the graft material powder may comprise the step of adsorbing bone morphogenic protein using a refrigerated centrifuge.

In some cases, the bone morphogenic protein may also be suspended in the solution. However, when the bone morphogenetic protein is adsorbed while rotating the bone morphogenetic protein at a high speed using a centrifuge, the bone morphogenetic protein can be prevented from being suspended in a solution, and thus the bone morphogenetic protein can be easily adsorbed onto the surface of the graft material powder or into the pores of the graft material powder. Only when the bone morphogenetic protein is adsorbed while rotating the bone morphogenetic protein at a high speed, the bone morphogenetic protein can be prevented from being resuspended after falling off from the graft material powder. If the bone morphogenic protein is rotated at a low speed, the bone morphogenic protein may be suspended and thus not easily adsorbed. Under high-speed rotation, the bone morphogenetic proteins can be rapidly adsorbed onto the surface of the graft material powder or into the pores of the graft material powder.

According to one embodiment of the present disclosure, the rotational speed of the refrigerated centrifuge may be 4,000rpm or greater. When a centrifuge is used to effect protein adsorption of bone morphology, the higher the rotation speed, the better the adsorption. For example, the rotational speed of the centrifuge may be 4,000rpm or more, and when this rotational speed range is satisfied, the bone morphogenetic protein can be prevented from being suspended in the solution.

According to one embodiment of the present disclosure, the step of adsorbing bone morphogenic protein using a refrigerated centrifuge can be performed at a cold temperature of 5 ℃ or less. Since the step of adsorbing bone morphogenetic proteins using a refrigerated centrifuge is performed at a cold temperature of 5 ℃ or less, it is possible to maximize the effect of adsorbing bone morphogenetic proteins onto the surface of the graft material powder or into the pores of the graft material powder by rotation while preventing the denaturation of thermolabile bone morphogenetic proteins by preventing the temperature of the solution from rising due to the rotation. The cold temperature may be a temperature at which the solution does not freeze. For example, the cold temperature may be 5 ℃ or less, and preferably, may be 0.5 ℃ to 1.5 ℃.

Thereafter, the graft material powder having the bone morphogenetic protein adsorbed thereon and the hydroxypropylmethylcellulose powder were mixed and stirred to form a viscous gel. The viscous gel thus formed can improve the adhesiveness of the graft material powder. For example, stirring may be performed using a mixer. When the graft material powder is stirred with hydroxypropylmethylcellulose in the form of powder, a product having a uniform quality can be obtained.

According to one embodiment of the present disclosure, the volume ratio between the graft material powder having the bone morphogenetic protein adsorbed thereon and the hydroxypropylmethylcellulose powder may be 1:0.2 to 1: 0.6. If the volume ratio of the graft material powder having the bone morphogenetic protein adsorbed thereon to the hydroxypropyl methylcellulose powder is greater than 1:0.2, it may be difficult to form a gel, and if the volume ratio of the graft material powder having the bone morphogenetic protein adsorbed thereon to the hydroxypropyl methylcellulose powder is less than 1:0.6, it may be difficult to form an effective bone graft composition because the volume of the gel is greater than the volume of the graft material powder. The volume ratio between the graft material powder having the bone morphogenetic protein adsorbed thereon and the hydroxypropyl methylcellulose powder may preferably be 1:0.25 to 1:0.35 in terms of the effect of the present disclosure.

Thereafter, the mixture of the graft material powder and the hydroxypropyl methylcellulose powder obtained by the mixing and stirring process was freeze-dried under vacuum to form a sponge-like structure comprising a plurality of pores. The sponge-like structure comprising a plurality of pores may also be formed by freeze-drying a mixture of graft material powder and hydroxypropylmethylcellulose powder obtained by a mixing and stirring process at a low temperature under vacuum.

The sponge-like structure including the porous structure may be formed by a freeze-drying process under vacuum. The gel may be absorbed into the graft material powder to form a sponge-like structure including a porous structure, and it is believed that the treatment under vacuum mainly contributes to the formation of the sponge-like structure including a porous structure.

According to one embodiment of the present disclosure, the method for preparing a bone graft composition may further comprise an encapsulation step.

According to one embodiment of the present disclosure, the method for preparing a bone graft composition may further include the step of placing the prepared bone graft composition including a sponge-like structure including a plurality of pores in a snap-on tube sized to be inserted into a syringe. When the method further includes the step of placing the composition in a snap tube having a size suitable for insertion into a syringe, the composition may have a size suitable for insertion into the syringe and thus may be directly inserted into the syringe without a separate process, so that the operation of the process for preparing the bone graft composition may be facilitated.

According to an embodiment of the present disclosure, the method for preparing a bone graft composition may further include the step of placing the bone graft composition including a sponge-like structure including a plurality of holes, which is placed in the overlap tube, in a syringe and sealing. When the bone graft composition is disposed in a syringe, ease of use can be ensured and the possibility of contamination that may occur during use is significantly reduced.

According to one embodiment of the present disclosure, the method for preparing a bone graft composition may further comprise the step of sterilizing the composition.

In one embodiment of the present disclosure, a bone graft composition comprising a sponge-like structure comprising a plurality of pores may be sterilized by ethylene oxide gas. For example, the concentration of ethylene oxide gas may be 450mg/L to 1,200 mg/L.

If the concentration of the ethylene oxide gas is less than 450mg/L, sterilization may be insufficient, and if the concentration of the ethylene oxide gas is greater than 1,200mg/L, denaturation of bone morphogenetic proteins may occur.

According to one embodiment of the present disclosure, a bone graft composition including a sponge-like structure including a plurality of pores may be sterilized by gamma ray irradiation. For example, the dose of gamma ray irradiation may be 10kGy to 25 kGy. If the dose of gamma irradiation is less than 10kGy, sterilization may be insufficient, and if the dose of gamma irradiation is more than 25kGy, denaturation of bone morphogenetic proteins may occur.

The bone graft composition prepared according to the above method should have a secured hydration ability to apply the composition to a human body, for example, to apply the composition to teeth. In other words, the bone graft composition should have critical osmotic properties. Having a certain permeability of the bone graft composition when mixed with another solution (e.g., saline) is an important factor in the ability of the bone graft material to be maintained at a constant concentration to maintain its shape and function. The property may be determined according to the content of HPMC and the like included in the bone graft composition and the condition of a solvent used to form a solution.

For example, in the case of applying a bone graft composition to a tooth, a dental practitioner applies the composition to the missing tooth after hydrating the bone graft composition provided as a powder or the like. If the HPMC contained in the bone graft composition flows out in a large amount (i.e., osmotic pressure of the hydrated composition increases) during the hydration process, the HPMC does not remain inside the bone graft material, and thus the bone graft material does not maintain its shape and does not cake, making it impossible to perform a medical procedure. For this reason, the bone graft composition should have permeability properties within a predetermined range so that it can maintain its shape without being affected by external conditions. In addition, if an external environment such as water or saliva is formed after the bone graft composition is applied to the missing tooth, a phenomenon in which the bone graft composition flows to the surroundings or falls off occurs, and thus a problem in that an external substance flows into the missing tooth may occur. For this reason, the bone graft composition should have a penetration property within a predetermined range, and in addition, the bone graft composition should have not only internal physical properties but also resistance to external conditions.

In fact, when the osmotic pressure of saline is set to 100% as a reference value, the saline should reach 104% to 112% of the osmotic pressure within 12 hours to 48 hours after the saline is added to the bone graft solution. If the osmotic pressure is more than 112%, HPMC may flow out during the hydration process, and thus the shape-retaining property of the bone graft composition and its adhesion to the bone defect may be significantly reduced, making it impossible to administer the bone graft composition. On the other hand, if the osmotic pressure is less than 104%, the bone graft composition may not be easily hydrated and thus may be difficult to be plastically deformed to the bone defect, thus adversely affecting bone regeneration.

Hereinafter, preferred examples will be given to aid understanding of the present disclosure. However, these examples are merely illustrative of the present disclosure and are not intended to limit the scope of the present disclosure as defined in the appended claims. In addition, it will be apparent to those skilled in the art that various changes and modifications of these examples are possible without departing from the scope and technical spirit of the present disclosure. In addition, it is to be understood that such changes and modifications are also within the scope of the appended claims.

Experimental examples

1. Measurement of osmotic pressure of brine

Osmotic pressure refers to the pressure applied to a semipermeable membrane when an osmotic phenomenon occurs, and is proportional to the difference in solution concentration. In the control experiment, the osmotic pressure of the saline was measured, and as shown in table 1 below, whether the osmotic pressure was changed with time was also measured. The osmotic pressure can be obtained by introducing a solution and measuring the value of the osmotic pressure by a pressure sensor connected to the outlet. The osmolality value of the saline measured in this control experiment was 286, which was set as 100% as a reference value.

2. Preparation of hydrated bone graft Material comprising hydroxypropyl methylcellulose (HPMC)

A hydrated bone graft material comprising HPMC was prepared by adding water to 1 part by weight (0.5g) of a bone graft material and 0.6 part by weight of Hydroxypropylmethylcellulose (HPMC). The amount of water added is 0.4 to 6 parts by weight per one part by weight of the mixture of the bone graft material and HPMC.

3. Measurement of osmotic pressure of saline added to hydrated bone graft material comprising hydroxypropyl methylcellulose (HPMC)

Each of the hydrated bone graft materials prepared in the above part 2 was placed in a 15mL conical tube, and 5mL of saline was added thereto. At each time point shown in table 1 below, 0.5mL of saline was taken and the osmotic pressure was measured. The measured osmolarity values are expressed as a percentage (%) relative to the reference values obtained in the control experiment of section 1 above.

TABLE 1

1 hour

3 hours

6 hours

12 hours

24 hours

48 hours

Control

100％

100％

100％

100％

100％

100％

0.4

100％

108％

112％

113％

115％

117％

0.5

100％

104％

109％

110％

111％

112％

0.6

100％

103％

107％

109％

110％

111％

0.8

100％

103％

106％

107％

108％

109％

1.0

100％

102％

103％

106％

107％

108％

1.2

100％

103％

104％

105％

106％

108％

1.5

100％

101％

102％

105％

107％

108％

2.0

100％

102％

102％

104％

106％

107％

3.0

100％

109％

115％

116％

117％

117％

4.0

100％

109％

116％

117％

118％

118％

6.0

100％

110％

114％

116％

118％

118％

As shown in table 1 above, the values obtained in the control experiment were set to 100% as reference values. This value is a value obtained by normalizing the osmotic pressure of pure brine measured at each time point.

As shown in table 1 above, it was confirmed that the osmotic pressure increased with time when the amount of added water (parts by weight) was constant. This indicates that the contact time and amount between the saline and the hydrated bone graft material comprising dissolved HPMC increases over time, thus increasing the osmotic pressure.

As shown in table 1 above, it was confirmed that the osmotic pressure increased as the amount of added water (parts by weight) increased at the same time point. This indicates that HPMC in the hydrated bone graft material comprising dissolved HPMC dissolves faster with increasing amount of added water, thus increasing osmotic pressure.

As shown in fig. 2, it was confirmed that, when the amount of added water was 0.4 parts by weight or less, the osmotic pressure was greatly increased in a short time, and thus the permeability was greatly changed, and even when the amount of added water was 3 parts by weight or more, the osmotic pressure was greatly increased in a short time, and thus the permeability was greatly changed. The fact that the osmotic pressure is greatly increased in a short time means that HPMC is not fused with the bone graft material but flows out, and thus the osmotic pressure is increased with the increase of the concentration, resulting in the decrease of the shape retentivity. Therefore, in order to optimize shape retention, the amount of water added is preferably set to 0.5 to 2 parts by weight based on 1 part by weight of the bone graft composition including HPMC.

When the amount of water added is 0.5 to 2 parts by weight based on 1 part by weight of the bone graft composition, the osmotic pressure of the saline may reach 104 to 112% in 12 to 48 hours. The function maintaining property of the bone graft composition is not sufficiently understood by the osmotic pressure value reached within 12 hours, and when the value is applied as it is, the demonstration of the functionality and stability of the composition as a bone graft material will be somewhat insufficient. Therefore, by examining the osmotic pressure reached within 12 hours to 48 hours, the functionality, osmotic properties, shape retention, etc. of the bone graft material can be determined. If the osmotic pressure within 48 hours exceeds 115% of the initial osmotic pressure of saline, it means that hydration of the bone graft composition comprising HPMC does not occur well. In this case, water is not sufficiently used as an additive, and thus the composition is difficult to use as a bone graft composition. If the osmotic pressure within 48 hours is close to 100%, it is not different from that of saline, and thus it is considered that the bone graft composition comprising Hydroxypropylmethylcellulose (HPMC) has little desired properties and functions. Therefore, it is preferable to prepare a bone graft composition to which saline reaches an osmotic pressure of 104% to 112% of that obtained in the control experiment within 12 hours to 48 hours after the addition thereof.

As described above, the bone graft composition forms an optimal osmotic pressure with another solution, and thus has excellent effects in activating bone formation, biocompatibility, and ease of use.