研發兒茶素改質之抗菌型牙科複合樹脂

Development of Bacteriostatic Epigallocatechin Gallate Modified Dental Composite Resin

蔡昀蓉(Yun-Rong Tsai)

指導教授 ： 姜昱至

國立臺灣大學/牙醫專業學院/臨床牙醫學研究所/碩士(2013年)

https://doi.org/10.6342/NTU.2013.10853

摘要

齲齒是經由反覆酸化、去礦化、再礦化最後形成窩洞；其中細菌形成細菌生物膜並造成副產物，使牙齒無機質溶解及有機質降解。兒茶素中之表沒食子兒茶酚沒食子酸（EGCG）為茶葉茶多酚裡面的主成分，具有多樣重要的生理作用如抗氧化、抗發炎、抗菌、抗酸蝕等。
光固化複合樹脂為目前主要用於齲齒窩洞填補的牙科填補材，但仍因其聚合收縮，導致填補物發生邊緣性滲漏，以及容易聚集細菌貼附而形成生物膜等特性，而難以抵抗繼發性齲齒的發生。
因此本研究目的為利用EGCG抑制主要致齲齒細菌—轉糖鏈球菌之能力，研發EGCG改質之牙科光固化複合樹脂填補材料，期能達抗菌防齲齒之功效。
本研究分三大部分進行，第一部分為基礎抗菌測定，包含轉糖鏈球菌生長曲線及EGCG之最小抑菌及殺菌濃度測定。
第二部分與台大高分子所合作開發抗菌型EGCG改質複合樹脂。針對EGCG成分作化學改質及接枝，以二異氰酸異佛爾酮（IPDI）及甲基丙烯酸-2-羥基乙酯（HEMA）反應形成先驅物，以異氰酸酯官能基（NCO）和表沒食子兒茶素沒食子酸酯上的羥基反應；以EGCG最小抑菌濃度250 μg/ml及最小殺菌濃度500 μg/ml為基準，將改質EGCG於所合成抗菌型複合樹脂之濃度分別為0 μg/ml、125 μg/ml、250 μg/ml、500 μg/ml、1500 μg/ml及3000 μg/ml。
第三部分為評估其抗菌效果與物理性質。研究結果顯示，市售不含抗菌成分之Estelite複合樹脂與不含EGCG之控制組樹脂生長細菌結果相當；改質EGCG濃度125 μg/ml、250 μg/ml之組別顯示之抗菌效果與宣稱可透過釋放氟離子達到抗菌效果之市售Beautifil樹脂相當；濃度提升到500 μg/ml以上，在電子顯微鏡及雷射共軛焦顯微鏡檢視下，轉糖鏈球菌的生長已相當稀少且活菌數量降為死菌1/2，顯示較市售產品更加優異的抗菌效果。
以紅外線光譜儀分析，顯示本實驗改質EGCG之添加濃度對複合樹脂之轉化率並無影響，且轉化率已高於市售Estelite樹脂。直徑抗拉伸強度試驗及表面微硬度測試之結果亦證實，改質EGCG之添加對這兩項性質並無影響，且數值皆達到與市售Estelite樹脂相當，證明其物理性質強度足以應用於臨床。
綜合以上結論，本研究建議以含改質EGCG 500 μg/ml為本填補材料之建議添加劑量。本研究建立一套標準化檢視牙科複合材料抗菌能力之實驗模式，而本實驗所研發之含EGCG改質之牙科光固化複合樹脂確實具有抵抗主要致齲齒細菌—轉糖鏈球菌之能力，且其物理性質及機械性質不下於市售產品之表現，故在臨床應用上對於預防繼發性齲齒發展應具有相當的潛力。

關鍵字

表沒食子兒茶酚沒食子酸 ； 兒茶素 ； 複合樹脂 ； 抗菌測試 ； 生物膜 ； 轉糖鏈球菌 ； 繼發性齲齒

並列摘要

Dental caries was a repeat process of acidification, demineralization, remiceralization and result in cavities at the end. Bacteria formed biofilms and produced acid byproducts, to cause dissolving of inorganic mineral and degradation of organic matrix of tooth structure. The epigallocatechin gallate（EGCG） was the main component of tea polyphenols in tea leaves, and has many important physiological functions such as anti-oxidation, anti-inflammation, anti-bacterial and anti-erosion.
Light-cured composite resin was the main dental filling material for caries cavity restoration nowadays. But because of its polymerization shrinkage to cause marginal leakage, and easily for biofilm formation by bacteria adhesion, it still didn’t have the resistant ability to secondary caries.
Therefore, this study aims to use the ability of inhibited the main cause bacteria of dental caries-Streptococcus mutans（S.mutans） of EGCG, we developed EGCG-modified light-cured composite resin dental filling material that has anti-bacterial and anti-caries effect.
This study carried out in three parts—Part 1: The basic anti-bacterial test, including the growth curve of S. mutans and the minimum inhibitory concentration（MIC） and minimal bactericidal concentration（MBC） of EGCG.
Part2: Development of EGCG modified dental composite resin, cooperation with the Institute of Polymer Science and Engineering of National Taiwan University. Modify EGCG by isocyanate functional group(NCO) from the oligomer of isophorone diisocyanate(IPDI) and methyl acrylate 2-hydroxyethyl methacrylate(HEMA), to chemical bond with hydroxyl group(OH) on EGCG. Base on the results of EGCG’s minimum inhibitory concentration: 250 μg/ml and the minimum bactericidal concentration: 500 μg/ml, the EGCG modified concentration for synthesising the antibaterial composite resin was decided of 0, 125, 250, 500, 1500 and 3000 μg/ml.
Part 3: To assese the antibacterial effects and physical properties of the composite resin. The results show that without EGCG, our control group composite resin is similar with commercial composite resin Estelite, which grow bacteria and fromed biofilm. The antibaterial effects of our EGCG modified composite resin with the concentraion 125 g/ml and 250 μg/ml are similar with the commercial composite resin Beautifil which claimed to have antibacterial effect by flouride releaseing. When the concentration of EGCG raised to 500 μg/ml, electron microscope and laser confocal microscope showed that the number of S. mutans decrease dramatically, and the number of viable bateria was less than half of the number of dead bacteria. , more excellent than the commercial product.
The results tested by Fourier transform infrared spectroscopy(FT-IR) showed that the conversion rate of composite resin was not influnced by the concentration of EGCG we added. And the conversion rate of our EGCG-modified composite resin was already higher then the commercial composite reisn Estelite. Diameter tensile strength test and surface microhardness test also confirmed that the concentration of modified EGCG had no effect on these two properties, and the values were similar or even higher than the commercial composite resin Estelite. These all showed that our EGCG-modified composie resin have good physical properties for clinical using.
Based on all the findings, our study suggested adding 500 μg/ml of modified EGCG into this composite resin. We established a standardized experimental mode of testing the antibateria effect of dental composite. The EGCG modified light curing dental composite resin are indeed have the ability to avoid adhesion and accumulation of the S. mutans, the principal cause bateria of dental caries. And its physical and mechanical properties are good enough when compared to the commercial composite resin. So we believed that this novel composite resin has considerable potential of clinical applications in the future.

並列關鍵字

Epigallocatechin gallate(EGCG) ； Catechin ； Composite resin ； Antibaterial tests ； Biofilm, Streptococcus mutans(S.mutans) ； Secondary caries

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