KR101984384B1 - Composition for Dental Restoration - Google Patents

Abstract

The present invention relates to a dental restorative composition comprising a powder containing zinc oxide and zirconia and a liquid emulsion, and is excellent in mechanical properties, cell compatibility and anti-inflammatory effect.

Description

Composition for Dental Restoration [0002]

The present invention relates to a dental restorative composition comprising a powder comprising zinc oxide and zirconia, and a liquid eugenol.

Zinc oxide-zeolite (ZOE) cement has been widely used for more than 100 years as a material for temporary restoration and bonding in dentistry because it is easy to handle, has good sealing, and has heat insulation and antibacterial properties. Also, temporary packing techniques using zinc oxide-eugenol cement incorporated dental sealants and zinc oxide-eugenol cements have been clinically used to treat dry sockets. Generally, the zinc oxide-eugenol cement is formed by mixing a zinc oxide powder and a liquid eugenol. At this time, the zinc-e nanolate chelate matrix phase is formed through an acid-base reaction, and unreacted zinc oxide acts as a reinforcing material.

However, such materials have poor mechanical properties, which can lead to failure of tooth restoration in patients with general dentition or strong occlusal force at the main occlusion point to which the material is applied. In addition, the cytotoxicity of zinc oxide-eugenol to cells originating from pulp tissue is a limit to utilization as a base material. In particular, zinc oxide-eugenol cements are problematic when applied to deep holes. Extracts of zinc oxide-eugenol cement in dentinal fluid can adversely affect pulp (stem) cells through open iv tubules during or after curing time.

In order to solve this problem, cement is known in which a specific additive is added to powder or liquid phase. For example, polymeric (resin) -molded zinc oxide-eugenol cements (e.g. IRM) and ethoxybenzoic acid-alumina-reinforced zinc oxide-eugenol cements have better mechanical properties and are clinically used Amin M. et al, Pediatr Dent 2016; 38: 317-24., Benz K et al., Aust Endod J 2016.) However, caution is still required when applying these to deep holes. Because of the large amount of zinc released during or after curing, these materials have an adverse effect on the survival rate of pulp (stem) cells through open dentinal tubules. Thereby reducing the regenerative capacity of the tooth-pulp complex. Therefore, it is necessary to solve the problems of zinc oxide-eugenol-based cement to improve the compatibility with pulp stem cells by reducing the release of zinc ions as well as mechanical properties.

Until now, there has been no research other than polymers, resins and alumina as additives for reinforcing zinc oxide-eugenol cement. In particular, zinc oxide-zeolite (ZOE) based cement incorporating zirconia (ZrO ₂ ) into powders for zinc oxide-eugenol is not known at all.

Under such circumstances, the inventors of the present invention have found that a dental restorative material in which zirconia is incorporated in powder form has an effect of improving not only physical properties but also excellent cell suitability as compared with conventional zinc oxide-eugenol cement, and further reducing the inflammatory reaction And it is very suitable as a dental restorative material.

One object of the present invention is to provide a dental restorative composition comprising a powder comprising zinc oxide and zirconia, and an eugenol.

Another object of the present invention is to provide a dental restoration method using the dental restoration composition.

It is another object of the present invention to provide a method for increasing the mechanical strength of a dental restoration composition comprising zinc oxide and eugenol.

It is still another object of the present invention to provide a method for reducing the cytotoxicity of a dental restoration composition comprising zinc oxide and eugenol.

It is still another object of the present invention to provide a method for manufacturing the dental restoration composition.

Another object of the present invention is to provide a kit for manufacturing a dental restoration packaged in a powder containing zinc oxide and zirconia, and in a liquid container for a liquid emulsion.

Hereinafter, the present invention will be described in more detail.

On the other hand, each description and embodiment disclosed herein can be applied to each other description and embodiment. That is, all combinations of the various elements disclosed herein are within the scope of the present invention. Further, the scope of the present invention can not be said to be limited by the following detailed description.

In addition, those of ordinary skill in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described in this application. Further, such equivalents are intended to be included in the present invention.

One aspect of the present invention to solve the above problems is to provide a dental restoration composition.

Specifically, the dental restorative composition comprises a powder comprising zinc oxide and zirconia, and a liquid eugenol.

The term " zinc oxide " as used herein is an inorganic compound having the formula ZnO. As the white powder, zinc oxide having a particle size of 1 nm or more and 100 占 퐉 or less can be used. Specifically, zinc oxide having a particle size of 1 nm or more and 100 nm or less can be used. If the size of the particles is smaller than this range, it may be difficult to uniformly disperse the particles in the composition due to the cohesive force between the particles. On the contrary, if it is larger than this range, the texture of the composition may decrease and it may be difficult to apply to teeth.

The zinc oxide can be prepared by various methods, and commercialized ones can be purchased and used. Since the zinc oxide-eugenol cement composition (ZOE) is commonly used as a dental material, it is possible to purchase zinc oxide powders and liquid emulsion which are sold for the purpose.

The term " zirconia " as used herein is an inorganic compound having the formula ZrO ₂ . Zirconia itself is used as a ceramic material and exhibits monoclinic form at room temperature. It is known that at a high temperature, a phase transition occurs in the form of tetragonal or cubic and a stable state can be formed. In the present invention, zirconia powder at room temperature can be used as an additive.

The zirconia powder can be produced by a solid phase method, a liquid phase method, or a vapor phase method, and any method may be used. Since zirconia is already used as a ceramic, commercialized zirconia can be purchased and used. Zirconia powder having a particle size of 1 nm or more and 100 占 퐉 or less can be used. In one embodiment of the present invention, zirconia having an average particle size of 5 mu m was used.

The dental restorative composition using zirconia as an additive in the present invention exhibits excellent mechanical properties as compared with the case where it is not included, does not cause toxicity in cells, and can minimize inflammatory reaction.

The zirconia may contain 1 to 25% by weight of the total powder. More specifically, it may contain 5 to 15% by weight of the total powder. More specifically, it may contain 15 to 20% by weight of the total powder. If the zirconia is contained in a larger amount than the above-mentioned content, the viscosity of the restoration material increases so that it becomes difficult to use it for the purpose of restoration. When the content of zirconia is less than the above content, the mechanical properties can not be improved to a desired extent or the cytotoxicity can not be alleviated to a desired extent. In one embodiment of the invention, zirconia is included by 5, 10, and 15 weight percent of the total powder.

The powder of the present invention may further comprise a polymer. The polymer may be any polymer that can be added to reinforce the zinc oxide-eugenol cement.

The polymer may be polyacrylates, polyolefins, polymethacrylates, or the like. More specifically, polyvinyl chloride (PVC), polyvinyl acetate (PVA), polystyrene, polymethyl acrylate (PMA), polymethyl methacrylate (PMMA) , Polyacrylic acid (PAA) and polyethylene, and polypropylene. However, the present invention is not limited thereto. In one embodiment of the present invention, a powder further comprising polymethyl methacrylate (PMMA) was used.

The term " eugenol " in the present invention is a transparent pale yellow essential oil extracted from clove oil, nutmeg, cinnamon, basil and bay leaves, and is a major component accounting for about 72 to 90% of the essential oil extracted from clove. Eugenol has the structure of the following formula (3) when R is hydrogen (H) in the structure of the above formula (1) and is also referred to as 4-allyl-2-methoxyphenol.

 [Chemical Formula 1]

Since the eugenol is clinically used for dentistry, commercially available products can be purchased and used. The eugenol can be extracted and purified from clove oil, nutmeg, cinnamon, basil and bay leaves by a method known in the art , And those chemically synthesized can be used. Since the zinc oxide-eugenol cement composition is commonly used as a dental material, it is possible to purchase zinc oxide powder and liquid emulsion which are sold for the purpose of use.

The eugenol may contain 90 wt% or more of the total liquid phase. Specifically, it may contain 95 wt% or more. More specifically, it may contain 98 wt% or more. In one embodiment of the present invention, a 100% liquid emulsion was used.

The powder and the liquid eugenol may be included in a ratio of 3 to 10 (g): 1 (mL). More specifically, it may be contained in a ratio of 4 to 6 (g): 1 (mL). If the proportion of the powder is smaller than the above ratio, the hardening time becomes long, which can make it difficult to use clinically. If the proportion of the powder is larger than the above ratio, mixing may become difficult. In one embodiment of the present invention, 5 g and 1 mL of powder and liquid eugenol, respectively, were used.

As used herein, the term " dentist " may refer to the use of teeth and their supporting tissues and mouths to prevent or treat disease or damage. The tooth may be enamel, dentin and cementum.

The compositions of the present invention are useful as restorative filling materials, preventive dental materials, pit and fissure sealants, temporary filling materials, and post- and core material, but is not limited thereto.

The composition of the present invention may contain other components such as polymers, ceramics, and metallic materials to the extent that physical properties and biological properties are not changed. Examples of the pigments include, but are not limited to, yellow pigments, navy blue and red iron oxide pigments, and titanium dioxide inorganic pigments.

The dental restorative composition of the present invention can be hardened after being filled in the pores of the teeth. The curing time can be increased as compared to a composition that does not incorporate zirconia.

The cured composition of the present invention incorporating zirconia may have excellent mechanical properties. In the experimental examples of the present invention, it was confirmed that the compressive strength, 3 contact bending strength, and Vickers hardness were increased in the specimens of the examples.

In addition, the cured compositions of the present invention may exhibit low cytotoxicity. In particular, the release of zinc ions is reduced and does not show cytotoxicity to pulp (stem) cells, so that it can be safely used. In the Experimental Example of the present invention, it was confirmed that the amount of zinc released in the test piece of the example was reduced and the hDPSC did not show cytotoxicity.

In addition, the cured compositions of the present invention may also exhibit an effect that may alleviate inflammation. In the experimental examples of the present invention, it was confirmed that the inflammation-related markers were reduced in the test pieces of the examples.

Another aspect of the present invention to solve the above problems is to provide a method of restoring a tooth using the dental restoration composition.

The dental restoration composition is as described above.

A method of providing a method of restoring a tooth includes filling a hole in the tooth with a dental restoration composition; And curing the composition.

The hole may refer to a portion of the teeth where the enamel or dentin has been corroded, for any reason, such as physical or chemical causes or decomposition by caries bacteria.

The cured composition can be softened by polishing with paper or the like.

The method can effectively restore teeth due to the excellent mechanical properties, cell fit, and anti-inflammatory effects of the restorative composition used.

Another aspect of the present invention to solve the above problems is to provide a method for increasing the mechanical strength of a dental restoration composition comprising zinc oxide and eugenol.

Another aspect of the present invention to solve the above problems is to provide a method for reducing cytotoxicity of a dental restoration composition comprising zinc oxide and eugenol.

Another aspect of the present invention to solve the above problems is to provide a method for increasing the anti-inflammatory activity of a dental restorative composition comprising zinc oxide and eugenol.

The dental restoration composition of the present invention is a dental restorative composition comprising zinc oxide and eugenol which further comprises zirconia powder in the zinc oxide powder to improve the mechanical strength of the dental restorative composition including zinc oxide and eugenol , Decrease cytotoxicity, and increase anti-inflammatory activity.

Another aspect of the present invention to solve the above problems is to provide a method of manufacturing the dental restoration composition.

The dental restoration composition is as described above.

Specifically, it may include mixing the powder containing zinc oxide and zirconia, and the liquid eugenol.

The powder containing zinc oxide and zirconia may be first mixed using a rolling ball. Mix for 12 hours or more, specifically 20 hours or more.

Thereafter, the powder and the liquid eugenol can be mixed at a ratio of 3 to 10 (g): 1 (mL) and 4 to 6 (g): 1 (mL).

Another aspect of the present invention for solving the above-mentioned problems is to provide a kit for manufacturing a dental restoration material packed in a separate container and powder containing zinc oxide and zirconia, and liquid emulsion play.

Specifically, the powder containing zinc oxide and zirconia, and the liquid emulsion are as described above.

The kit may be prepared by preparing a dental restoration composition in the form of a paste by mixing the powder phase with a liquid phase immediately before use, and then injecting the paste into a desired treatment site.

The dental restorative composition of the present invention comprising powder of zinc oxide and zirconia, and liquid eugenol has excellent mechanical properties. In addition, the amount of zinc released is low, the cell suitability is excellent, and the inflammation of the teeth can be alleviated.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph showing curing time and mechanical properties of Examples 1 to 6 and Comparative Examples 1 and 2. FIG. 1A is a diagram showing a curing time (n = 5). 1B is a diagram showing the compressive strength (n = 5) measured by the ISO standard method (ISO 3107). Fig. 1C is a diagram showing a three-contact bending strength (n = 10). Fig. FIG. 1D is a diagram showing Vickers hardness (n = 5). FIG.
Fig. 2 shows the morphologies and surfaces of Examples 1 to 6 and Comparative Examples 1 and 2; Fig. FIG. 2A is a graph showing the results of scanning electron microscopy to observe the surface morphology of the composition after 24 hours of curing the example and comparative compositions. FIG. 2B is a view for confirming the amount of released zinc and Zr ions according to the energy dispersive X-ray spectroscopy. FIG. 2C is a graph showing a Zr signal measured according to element mapping. FIG.
FIG. 3 is a graph showing the concentrations of zinc ions and eugenol released from the extracts in the SM medium of the specimens of Examples 1 to 6 and Comparative Examples 1 and 2. FIG. Figure 3A shows the concentration of zinc ions and Figure 3B shows the concentration of eugenol.
Fig. 4 shows the cell suitability of Examples 1 to 3 and Comparative Example 1. Fig. FIG. 4A shows cell viability of hDPSCs under ZnCl2, eugenol, and mixed conditions thereof. FIG. Figures 4B-D show hDPSCs cell viability in extracts of SM medium (concentration 50%, 25%, and 12.5%, respectively) of Examples 1 to 3 and Comparative Example 1.
Fig. 5 is a diagram for the continuous anti-inflammatory effect of Examples 1 to 3 and Comparative Example. Fig.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are intended to illustrate the present invention, but the scope of the present invention is not limited thereto.

Manufacturing example . Dentistry Restorative  Method for producing a composition

Example 1

A commercially available zinc oxide eugenol cement (ZOE) consisting of zinc oxide (ZnO) powder and 100% liquid effervescent was purchased (Kemdent, Purton, UK; Powder Production Number: 23705, 22417). Zirconia (ZrO2) was incorporated in an amount of 5% by weight based on the total amount of the powders. After mixing the materials mechanically for 24 hours using a rolling ball, the liquid eugenol was added to the powder in a ratio of 1 (ml): 5 (g). The dental restorative compositions were prepared by mixing according to the manufacturer's instructions.

Examples 2 and 3

The dental restoration composition was prepared in the same manner as in Example 1 except that the mixed injection of zirconia was made to be 10 and 15% by weight with respect to the total amount of powder, respectively.

Comparative Example 1

A dental restoration composition was prepared in the same manner as in Example 1 except that zirconia was not incorporated.

Examples 4 to 6

A commercially available reinforced zinc oxide eugenol cement (IRM) was purchased (Dentsply, York, PA, USA, Powder Production Number: 1407283, Liquid Phase Number: 1407283) in the same manner as Examples 1 to 3 , And zirconia were incorporated at 5, 10, and 20 wt%, respectively, based on the total amount of powder, to prepare a dental restorative composition.

Comparative Example 2

A dental restoration composition was prepared in the same manner as in Example 4 except that no zirconia was incorporated.

The above Examples 1 to 6 and Comparative Examples 1 and 2 were appropriately stored under the manufacturer's recommended conditions before the experiment. Unless otherwise specified, in the experiments described below, polishing was carried out with a maximum of 1200 grit SiC paper before use.

Examples 1 to 3 and Comparative Example 1 are summarized in Table 1 below.

Powder (% by weight) Liquid phase Mixing ratio ZnO ZrO2 Comparative Example 1 100% 0% Eugenol 5: 1 Example 1 95% 5% Eugenol 5: 1 Example 2 90% 10% Eugenol 5: 1 Example 3 85% 15% Eugenol 5: 1

Experimental Example  1. Curing time measurement

1-1. Test Methods

The curing times of the compositions of Examples 1 to 6 and Comparative Examples 1 and 2 were determined according to the ISO standard method for zinc oxide eugenol cements (ISO 3107, Dentistry-Zinc oxide / eugenol cements and zinc oxide / non-eugenol cements Switzerland: International Standardization Organization; 2011). Briefly, after 60 seconds from the start of mixing, the composition was filled into a mold (φ = 10 mm, d = 2 mm) and placed in a metal block. During the measurement period, the composition was stored at a temperature of 37 DEG C and a humidity of 95%. A flat indentor needle (400 g, ϕ = 1 mm) was placed on the composition surface at intervals of 15 seconds from 8 minutes after the start of the mixing. Curing time was recorded if the needle did not fully penetrate to the depth of 2 mm of the cement (n = 5). A clean needle tip was maintained during the measurement.

1-2. result

The cure time of Comparative Example 1 was about 9 minutes, which was less than the maximum cure time of 2 type zinc oxide eugenol cement in the ISO standard for basic and temporary restoration (Figure 1A). However, the curing time increased significantly with the incorporation of zirconia, and reached 17 minutes in Example 3 with 20% addition. Likewise, even with the use of reinforced zinc oxide eugenol cement, the curing time increased from 5.5 minutes to 7.5 minutes with incorporation of zirconia (FIG. 1A).

Experimental Example  2. Mechanical properties

2-1. Test Methods

(1) Compressive strength

The compressive strengths of the materials obtained by curing the compositions of Examples 1 to 6 and Comparative Examples 1 and 2, respectively, were measured according to ISO 3107 described above. Briefly, after completing the mixing of the composition and filling it into a cylinder mold (φ = 4 mm, h = 6 mm) within one minute, the mold was heated to 37 ° C. and 95% Humidity < / RTI > cabinet for 1 hour. The surface of the specimen was then polished and flattened and a suitable specimen (no defect or broken edge) was selected and immersed in distilled water at 37 ° C for an additional 24 hours before the compression test. The prepared specimens were placed on an Instron 5966 machine (MA, USA) for compression testing. The compressive strength was determined at a crosshead speed of 1.0 mm / min (n = 10). The compressive strength (MPa) was calculated by dividing the maximum application force (N) by the area of the specimen (mm ² ).

(2) 3 contact bending strength

The contact bending strengths of the specimens obtained by curing the compositions of Examples 1 to 6 and Comparative Examples 1 and 2, respectively, were measured using curved bar specimens made of stainless steel according to ISO 9917-1 (ISO 9917-1 Dentistry - water-based cements - part 1: powder / liquid acid-base cements. Switzerland: International Standardization Organization; 2007). The specimens were prepared under the same conditions as those used in the compressive strength test. Unblemished bar specimens without any defects or broken edges were prepared for the 3-contact bend test using a support spacing of 20 mm. The 3-contact bending strength was determined at a crosshead speed of 1.0 mm / min (n = 10) using the general purpose tester. The 3-point bending strength (σ) was calculated using the following formula:

σ = 3Fl / 2bh ²

(Where F is the maximum stress determined by the universal testing machine, l, b, and h are the span length, width, and height of the test specimen, respectively)

(3) Vickers hardness

The specimens obtained by hardening each of the compositions of Examples 1 to 6 and Comparative Examples 1 and 2 were stored at a humidity of 95% and a temperature of 37 캜 for 24 hours, and their Vickers hardnesses were measured. The surface of the cement was softened through contact with a polyethylene film (Printec, Daegu, Korea), and then Vickers hardness (HM-221, Mitutoyo, Tokyo, Japan) ). ≪ / RTI > Five specimens were measured (n = 5) per group of Examples and Comparative Examples, and the average value was taken as a representative value.

1-2. result

As the content of zirconia increased, compressive strength, 3-contact bending strength, and Vickers hardness gradually increased (see FIG. 1B-D, Comparative Example 1 <Example 1? Example 2 <Example 3). The compressive strength of Comparative Example 1 was 22.0 ± 3.7 MPa, the compressive strength of Example 3 was 33.8 ± 3.0 MPa, and the compressive strength was increased to 45% of Comparative Example 1 with zirconia addition (FIG. 1B). The contact bending strength of Comparative Example 1 was 3.4 ± 0.5 MPa, the contact bending strength of Example 3 was 8.5 ± 1.5 MPa, and the contact bending strength was significantly increased to 250% as compared with Comparative Example 1 by adding zirconia 1C). In addition, the Vickers hardness of Comparative Example 1 was 12.2 ± 0.9 HV, the Vickers hardness of Example 3 was 21.4 ± 1.0 HV, and the Vickers hardness increased to 75% as compared with Comparative Example 1 by adding zirconia (FIG. 1D).

Similarly, the compressive strength, the three-contact bending strength, and the Vickers hardness of Examples 4 to 6 using the reinforced zinc oxide eugenol cement (IRM) were found to be ~ 45%, ~ 130% And ~ 100% (see Figure 1B-D, P < 0.05).

Experimental Example  3. Surface properties

3-1. Test Methods

Disc samples (Φ = 10 mm, d = 2 mm) obtained by curing the compositions of Examples 1 to 6 and Comparative Examples 1 and 2 were polished using SiC paper (maximum 1200 grit). The surface morphology and composition were measured with a scanning electron microscope (SEM, Sigma 500, ZEISS, Jena, Germany) and a 15 kV energy dispersive X-ray spectroscopy (EDS, Ultradry, Thermo fisher, Waltham, MA, USA) Respectively. Elemental mapping was performed to investigate whether zirconia was uniformly dispersed in cement. The surface roughness (Ra) was measured according to the detailed procedure (SJ-400, Mitutoyo, Japan) at a rate of 0.5 mm / s with 4.0 mm scan (n = 5).

3-2. result

The Ra values of Comparative Example 1 and Examples 1 to 3 were found to be 0.23 ± 0.04, 0.35 ± 0.03, 0.48 ± 0.03, and 0.79 ± 0.09 μm, respectively. Likewise, the Ra values of Comparative Examples 2 and 4 to 6 were 0.35 ± 0.03 μm, 0.43 ± 0.04 μm, 0.64 ± 0.05 μm, and 0.96 ± 0.17 μm, respectively, which increased by 3 times with zirconia inclusion (p < 0.05). SEM images confirmed that the surface morphology of Examples 1 to 3 varied with the incorporation of zirconia when polishing with a maximum of 1200 grit of SiC paper (FIG. 2A). The composition of Comparative Example 1, Examples 1 to 3 was confirmed by EDS which is appropriate for the well-specified composition of each group (Fig. 2B). According to the elemental surface analysis, the Zr signal gradually increased with the incorporation of zirconia. In contrast, almost no background signal was detected in Comparative Example 1 (Fig. 2C).

From these results, it can be seen that the examples increase the mechanical properties due to the interaction with the ends of the cracks or the bridging effect of the cracks during the formation of the cracks due to the incorporation of zirconia .

Experimental Example  4. Releases of ions and chemicals

4-1. Test Methods

First, the specimens obtained by curing the compositions of Examples 1 to 6 and Comparative Examples 1 and 2, respectively, were sterilized with ethylene oxide (EO) gas using a Person EO50 system (Persson Medical, Gunma, Kyonggi-do). After disc-shaped specimens were cultured for 24 hours in a cell culture medium (3 cm ² / mL; composition described in Experimental Example 5), specimens (Φ = 10 mm, d (ICP-AES, Optima 4300 DV, PerkinElmer, n = 3) was used to analyze the Zn or Zr ions emitted from the sample. (GC-MS, Agilent, Santa Clara, Calif.) According to known protocols to measure the chemicals (eugenol and derivatives thereof) contained in the extract of the specimen cultured in the culture medium for 24 hours. Were used. Briefly, extracts of the specimens were lyophilized for 24 hours and volumes of ethanol equal to that of conventional extracts were added before performing GC-MS measurements.

4-2. result

Overall, the zinc ions were released in both the Examples and Comparative Examples, while the release of Zr ions was not detected (<0.1 ppm, FIG. 3A). Between 10 minutes and 24 hours after the start of mixing, Comparative Example 1 emitted Zn ions by 37.5 +/- 3.3 ppm, whereas Examples 2 and 3 released significantly less Zn (28.5 3.5 and 21.8 1.8 ppm, P &Lt; 0.05). The amount of Zn ions released between 17 minutes and 24 hours was 30.5 ± 2.5 ppm for Comparative Example 1, 18.9 ± 1.5 ppm for Example 3, and less than the amount of Zn ions released from 10 minutes to 24 hours (* 3A, P <0.05) . The order of Zn ion release was similar to that of 10 min to 24 h (P <0.05).

Only eugenol was detected in potentially releasable chemicals (eg, eugenol and zinc acetate) (Figure 3B). From 10 minutes to 24 hours after the start of the mixing, Comparative Example 1 released 21.3 1.5 ppm of eugenol but Examples 1 through 3 released at significantly lower concentrations (18.8 1.8, 15.8 1.1 and 13.4 1.51.1 ppm ). Significantly lower emissions between 17 minutes and 24 hours (* * 3B, P <0.05) compared to emissions between 10 minutes and 24 hours; The order of eugenol doses released between 17 minutes and 24 hours was similar to the order of release from 10 minutes to 24 hours (P <0.05).

Experimental Example  5. Cytotoxicity test

5-1. Test Methods

The first culture hDPSC (Human Dental Pulp Stem Cells) obtained from the third molar, which was not corroded, was used in low-pass (less than 10) in vitro experiments (the experimental protocol is the Institutional Review Board (H-1407/009/004) Approved by Supplemented with 10% fetal bovine serum (Gibco, Waltham, MA, USA), 1% penicillin / streptomycin (Invitrogen, Carlsbad, CA, USA), 2 mM GlutaMAX (Gibco) and 0.1 mM L-ascorbic acid The cells were cultured in an alpha-minimum essential medium (α-MEM) medium, which was kept at 37 ° C and humidified atmosphere containing 5% CO ₂ . This medium is hereinafter referred to as supplemented medium (SM).

The cytotoxicity test was performed according to ISO 10093-5 (ISO 10993-5 Biological evaluation of medical devices - part 5: tests for in vitro cytotoxicity. Switzerland: International Standardization Organization; 2009.). hDPSCs (1 x 10 ⁴ cells / mL) were inoculated into 96-well plates and cultured for 24 hours. After that, 50 mu L of SM medium and 50 mu L of SM medium containing 50%, 25% or 12.5% of the extract were added and cultured for 24 hours. The SM medium containing the above-mentioned sample extract was prepared by dissolving the disc specimen (Φ = 10 mm, d = 2 mm) obtained by curing the compositions of Examples 1 to 6 and Comparative Examples 1 and 2 according to ISO 10993-12 and SM medium ² / mL, and cultured for 24 hours using a shaking incubator (37 ° C, 120 rpm). 100 [mu] L of SM was used as a negative control (100% cell viability) and phenol (1%, Sigma) was used as a positive control (~ 0% cell viability). Cell viability (n = 6) was measured using a spectrophotometer (SpectraMax M2e, Molecular Devices, 490 nm) using the MTS assay (CellTiter 96, Promega, Madison, WI, USz) after 24 h of symbiotic incubation. Survival and death cells of 50% and 25% specimen extract treated groups were identified using kits (Molecular Probes, Eugene, Ore, USA) to identify cytotoxic results and the number of viable (green) and dead Images were taken using a confocal laser microscope (LSM 700, Carl Zeiss, Oberkochen, Germany).

All analyzes were carried out three times independently and representative values were noted.

5-2. result

Cytotoxicity of the test extract in cell culture medium (SM) was determined using WST assay. First, and (Fig. 4A) determines the Zn ^{2 +,} which results in 50% cell viability compared to the control (ZnCl ₂ derived) and the effective concentration of eugenol (i.e., EC50) (horizontal dashed line in Fig. 4). Within the irradiated detection range (~ 40 ppm), the EC50 values of Zn ion and eugenol were 9.1 ppm and undetermined (40 ppm or more), respectively. The cell viability of hDPSCs incubated with 40 ppm of eugenol and a certain concentration of Zn ions was not different from that of cells treated with Zn ion only.

When the same undiluted specimen extract was added to hDPSCs in SM medium, the concentration of specimen extract in the whole medium was 50%. At these 50% concentration of sample extracts, at 10 min and 17 min, Comparative Example 1 and Examples 1 and 2 showed very low levels of cell viability (less than 10%) as compared to the control (Figure 4B) (P < 0.05). In contrast, at 10 minutes (13.1 ± 4.3%) and 17 minutes (35.6 ± 4.3%), Example 3 showed significantly increased cell survival (P <0.05) compared to the other groups. A similar level of cell viability (about 100%) was observed in all of the examples (showing significantly lower cell viability than the control group), except for Comparative Example 1 at 25% extraction conditions (Figure 4C) 2.3% at 10 minutes, 88.3 ± 5.6% at 17 minutes, P <0.05). At the extraction conditions of 12.5% (Fig. 4D), the same cell viability was observed in Comparative Example 1, Examples 1 to 3 (P > 0.05).

The cytotoxic results were confirmed through the images of living cells and dead cells observed by confocal microscopy (Fig. 4E). A large number of dead cells (red) and a few viable cells (green) were observed at 10 and 17 minutes in the 50% sample extracts of Examples 1 and 2 and Comparative Example 1 (Examples 1 and 2 are not separately shown). In contrast, Example 3 showed more viable cells at the start of culture (10 min < 17 min) and correlated with the WST results.

In the 25% extract, Comparative Example 1 showed more viable cells than the control (Fig. 4E) in which the Comparative Example 1 was cultured for 10 minutes in the 50% extract according to the incubation start time (10 minutes < 17 minutes). In contrast, Examples 1 to 3 showed a similar amount of viable cells compared to the control group (Examples 1 and 2 are not separately shown).

Experimental Example  6. Anti-inflammatory  exam

6-1. Test Methods

HPSCs treated with LPS were incubated with the 25% sample extracts of Examples 1-3 or other concentrations of eugenol (1.25-5 ppm) for 24 hours. The concentration of the eugenol is within a range that can be released from the sample extract.

To confirm anti-inflammatory effects, the levels of IL-1β, IL-6 and IL-8 expression of normal and inflammatory hDPSCs were assessed by reverse transcriptase polymerase chain reaction (RT-PCR) and quantitative PCR (qPCR) . RT-PCR and qPCR were performed according to the manufacturer's protocol. Briefly, total RNA was extracted using an RNA purification kit (GeneAll, Ribospin, Seoul, Korea) and used with AccuPower RT Premix (Bioneer, Daejeon, Korea) and 2720 gene amplifiers (Life Technologies, Foster City, 2 μg of total RNA was reverse transcribed with cDNA. qPCR experiments were performed using RealAmp SYBR qPCR master mix (GeneAll) and RT-PCR system (StepOnePlus, Applied Biosystems) according to the manufacturer's instructions. The primer sequences used are listed in Table 2 below.

origin gene The forward primer sequence (5'-3 ') The reverse primer sequence (3'-5 ') SEQ ID NO: 1 Person β-actin AGG ATG CAG AAG GAG ATC ACT G ATA CTC CTG CTT GCT GAT CCA C SEQ ID NO: 2 Person IL-1? GGC AGA AAG GGA ACA GAA AGG AGT GAG TAG GAG AGG TGA GAG AGG SEQ ID NO: 3 Person IL-6 CTG GCA GAA AAC AAC CTG AAC ATG ATT TTC ACC AGG CAA GTC SEQ ID NO: 4 Person IL-8 CTA GGA CAA GAG CCA GGA AG AGT GTG GTC CAC TCT CAA TC

Samples were normalized with glycerol aldehyde-3-phosphate dehydrogenase (GAPDH) and the relative multiples of expression were automatically calculated using the 2 ^{- ΔΔCt} values for non-stimulated hDPSC by the StepOne software program v2.3 [8]. A representative mean (n = 4) with a standard deviation (SD) was recorded after three independent experiments.

3.4. Anti-inflammatory effect

MRNA expression levels of inflammatory markers (IL-1β, IL-6 and IL-8) in LPS-treated hDPSC were investigated after incubation with 25% sample extract for 17 minutes. Cell survival was ~ 90% in all groups. LPS stained hDPSCs were detected in negative control (IL-1β (7.5 ± 1.5), IL-6 (4.2 ± 0.5) and IL-1β (1.0 ± 0.2), IL- 0.2). When compared with LPS-treated hDPSCs containing no extracts, the addition of the extract of the Example significantly reduced the levels of mRNA expression of all inflammatory markers (IL-1β, 6 and 8) 1 were 1.2 ± 0.3, 1.4 ± 0.2, 1.5 ± 0.3, and 2.1 ± 0.2, respectively, for the reduced levels of Comparative Example 1, Example 1, Example 2, and Example 3. Normal noninflammatory hDPSC The levels of mRNA expression of the three inflammatory markers were significantly reduced compared to mRNA expression levels of the negative control (1, no LPS treatment, P <0.05); IL-6 in Example 3 IL-8 was the only exceptional increase. On the other hand, through direct treatment of eugenol with LPS-treated hDPSCs, mRNA expression levels of inflammatory markers were determined in a concentration-dependent manner (E1.25 < E2.5 < E5, P < 0.05).

From the above description, it will be understood by those skilled in the art that the present invention may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. In this regard, it should be understood that the above-described embodiments are to be considered in all respects as illustrative and not restrictive. The scope of the present invention should be construed as being included in the scope of the present invention without departing from the scope of the present invention as defined by the appended claims.

<110> Dankook University Cheonan Campus Industry Academic Cooperation Foundation <120> Composition for Dental Restoration <130> KPA170534-KR <160> 8 <170> KoPatentin 3.0 <210> 1 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> forward primer of beta-actin <400> 1 aggatgcaga aggagatcac tg 22 <210> 2 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> reverse primer of beta-actin <400> 2 atactcctgc ttgctgatcc ac 22 <210> 3 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> forward primer of IL-1beta <400> 3 ggcagaaagg gaacagaaag g 21 <210> 4 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> reverse primer of IL-1beta <400> 4 agtgagtagg agaggtgaga gagg 24 <210> 5 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> forward primer of IL-6 <400> 5 ctggcagaaa acaacctgaa c 21 <210> 6 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of IL-6 <400> 6 atgattttca ccaggcaagt c 21 <210> 7 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> forward primer of IL-8 <400> 7 ctaggacaag agccaggaag 20 <210> 8 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of IL-8 <400> 8 agtgtggtcc actctcaatc 20

Claims (13)

A powder mixture of zinc oxide powder and zirconia powder, and a liquid eugenol. 2. The dental restorative composition of claim 1, wherein the zirconia is from 1 to 25 weight percent of the total powder mixture. The dental restorative composition of claim 1, wherein the zirconia is 5-15 wt% of the total powder mixture. The dental restoration composition of claim 1, wherein the eugenol is at least 90% by weight of the total liquid. The dental restoration composition according to claim 1, wherein the powder mixture and the liquid emulsion are included in a ratio of 3 to 10 (g): 1 (mL). The dental restoration composition according to claim 1, wherein a polymer is added to the powder mixture. The method of claim 6, wherein the polymer is selected from the group consisting of polyvinyl chloride (PVC), polyvinyl acetate (PVA), polystyrene, polymethyl acrylate (PMA), polymethyl methacrylate wherein the composition is at least one selected from the group consisting of polymethyl methacrylate (PMMA), polyacrylic acid (PAA), polyethylene, and polypropylene. 7. The dental restorative composition of claim 6, wherein the polymer is 20 to 40 weight percent of the total powder. delete A powder mixture of zinc oxide powder and zirconia powder; And mixing the liquid effervescent powder with the liquid effervescent powder. The method of increasing the mechanical strength of a dental restorative composition comprising zinc oxide and eugenol. A powder mixture of zinc oxide powder and zirconia powder; And mixing the liquid eugenol with the dental restorative composition. The method of reducing the cytotoxicity of a dental restorative composition comprising zinc oxide and eugenol. A powder mixture of zinc oxide powder and zirconia powder; And mixing the liquid eugenol. A powder mixture of zinc oxide powder and zirconia powder, and a kit for preparing a dental restoration wrapped in a liquid emulsion play individual container.

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