變異鏈球菌 ( Streptococcus mutans ) 是造成口腔疾病的主要原因，此菌體藉由酶的作用 ( enzymatic action ) 將口中碳水化合物轉化成高黏滯性的胞外基質 ( extracellular polymeric substance, EPS )。EPS 由多醣體 ( Polysaccharides ) 組成，使變異鏈球菌附著於牙齒上之後，堆疊而形成生物膜。EPS 的結構具有許多孔洞及孔隙，能容納大量水分，使生物膜內的水分含量佔整體的 97 %，能夠幫助細菌運輸養分及廢物。S. mutans 生物膜對牙齦有許多不良影響，小則蛀牙，大則患上牙周病。形成生物膜的 S. mutans 由於有外層的 EPS 保護，許多抗菌劑無法直接對細菌本體作用而效果大大減低，故想要將 S. mutans 的活性消除，必須先破壞EPS或是找尋不被 EPS 阻擋的殺菌劑。
本實驗使用培養兩天的生物膜，分別以紫外光、微電漿、次氯酸水溶液以及酒精，以不同時間做處理。所使用的分析方法主要為原子力顯微鏡 ( atomic force microscope, AFM )，搭配電化學阻抗 ( electrochemical impedance spectroscope, EIS )、螢光染色等量測，藉由觀察生物膜之表面形貌、機械性質、電學性質及存活率，得到不同殺菌方法下，各種性質的變化。機械性質方面，彈性模數、黏滯力、形變量以及能量耗散分別對應於生物膜的彈性、EPS 含量、硬度 ( stiff ) 以及水分含量；而阻抗量測結果則能夠與水分多寡相對應。我們藉由探討以上數值的變化推測生物膜中 EPS 對不同刺激的反應。另外，使用 AFM 的峰力定量奈米機械性質測量模式 ( peak force quantitative nanomechanical property mapping, peak force QNM )，我們能夠在細菌表面形貌不產生改變時，得知生物膜中 EPS 的變化。藉由電性與螢光染色結果的輔助，我們的論點能更為穩固。
本實驗中利用 AFM的方法可在不傷害細菌的情況下量測生物膜機械性質，得知生物膜中 EPS 的變化。從結果中我們發現，生物膜對紫外光、微電漿、HOCl 及酒精的反應皆不一樣。紫外光能使 EPS 被分解而減少，卻不至於將結構瓦解，因此水分仍可被保留於生物膜中，而此時紫外光已可穿透過 EPS，消滅外層的 S. mutams 活性；EPS 結構被微電漿所產生的反應性物質攻擊而瓦解塌陷，而反應性物質也會因為 EPS 結構豬的孔洞以及孔洞內水分的散失而進入倒生物膜內部，作用於變異鏈球菌；在 HOCl 水溶液中，free chlorine 會與 EPS 反應，使得生物膜中的水分流失，EPS 結構強度下降，此時的 free chlorine 卻已被消耗不少，因此活性消失的 S. mutans 並不多；酒精對生物膜的影響在於消除 EPS 結構中的水分，導致 EPS 結構缺乏支撐而些微塌陷。
本實驗所使用的殺菌方法若欲將生物膜細菌全數消滅，則必須將各種方法交錯使用，或是將處理時間延長。然而，每一種殺菌劑的處理時間都有限制。在未來，期望能研究出有效率的方法，將此方法應用於醫療或工業，除了能治療疾病外，也能改善衛生問題。

In this study, we investigated the physiological status of Streptococcus mutans and extracellular polymeric substance (EPS) under different environmental stress factors such as UV light, micro-plasma, hypochlorous acid and alcohol by atomic force microscopy, electrochemical spectroscopy, and widefield microscopy. The effects of stressed environment were characterized in morphology, mechanical properties, bio-electrochemistry, and viability.
Based on the analysis, we infer that EPS can be damaged by UV light, micro-plasma, hypochlorous acid at various treatment levels. The disruption of EPS structure causes biofilm to lose the ability to transport nutrients and waste products. In our results, UV light, HOCl and plasma have to first disrupt EPS then attack bacteria. UV light reduce EPS and then penetrate into S. mutans, which inactivate S. mutans. The reactive species of plasma can damage the structure of EPS directly. After the free chlorine in HOCl solution react with EPS, EPS and the free chlorine are both consumed. Alcohol can remove most of the water in biofilm, and let the structure of EPS collapse slightly.
In the future, we can either mix each disinfectant to inactivate entire biofilm, or extend the treat time. We expect that these methods can be applied to medical treatments.

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