- 學位論文
可攜式微生物感測器：工作電壓對於不同固定菌屬電極之最佳化探討
Portable Microbial Biosensor : Optimization of Working Voltage for Varied Bacteria-Attached Electrode
摘要
摘要 本研究針對實驗室先前開發之微生物感測系統，其具有內建特殊算數程式，加以改良並使之最佳化以檢測廢水毒性。使用三種不同菌屬之菌株：枯草桿菌(Bacillus subtilis)、大腸桿菌（Escherichia coli）及惡臭假單胞菌（Pseudomonas putida）作為感測系統之生物電化學反應的感測元件。微生物經分批培養後以不同的溫度、pH值及工作電壓測試，試圖找出其最佳的微生物工作環境，並由攝氧率、培養皿計數法、混濁度法及顯微鏡檢法作菌數及活性判別。擬取其最佳培養組合的菌液做為附著於微生物感測器之電極試片的菌種。有毒水樣會抑制微生物對氧的攝取，因此產生的微小電流隨配製毒性劑量而改變。利用氧的消耗量轉換為電化學或電荷轉移效應，以兩者關係來推斷該水樣的毒性。本研究以提供不同工作電壓於電極試片上，使增進廢水毒性相對於電壓輸出的判別。量測之電壓與時間的特性圖，以三種數學計算模式表示而得到特定的抑制率，取與溶氧儀之抑制率最相近的數值，並取偵測時間為最迅速的該計算模式，提供一個具充分運算的內建辨識處理系統。分析結果顯示：在最佳電極試片之重活化時間及微生物的生物活性下，各植菌試片依不同菌屬微生物之最佳化工作電壓，即氧化還原反應速率快與菌體攝氧率最大，分別為：枯草桿菌0.85 V，大腸桿菌0.65 V及0.70 V、及惡臭假單胞菌0.70 V及0.75 V。以對酚的溶液為參考，三種不同菌屬的可偵測濃度範圍分別為：枯草桿菌在400-3200 mgl-1之間，大腸桿菌在200-1600 mgl-1之間，惡臭假單胞菌在400-2400 mgl-1之間。除此之外，運算模式對於判別酚的毒性，對照於溶氧儀所得到的結果有顯著的差異；以最佳的模式運算時，其相關係數分別為：枯草桿菌0.951、大腸桿菌0.903、及惡臭假單胞菌0.921。
並列摘要
Abstract In this study, our previous microbial sensing system with a built-in special processing algorithm has been upgraded and further developed for detecting wastewater’s toxicity. Bacillus subtilis, Escherichia coli and Pseudomonas putida were utilized as sensing components of the system to initiate biological and subsequent electrochemical reactions. The bacteria were incubated in batch and tested under different temperatures, pH values and working voltages. We tried to find out the optimized working environment of such bacteria. Bacterial numbers and their bioactivities were estimated using the oxygen uptake rate of bacteria, the plate count method、the turbidimetric method and the microscopic count method, respectively. The bacterial solutions incubated under such optimized combination were then attached onto the electrodes of the system. The infinitesimal currents were correlated with the variation of the defined dosages of toxicity tested on the bacteria-attached electrodes. The addition of toxic substances can inhibit oxygen uptake rate of the bacteria, which leads to induce electrochemical or charge transfer effects. Thereafter, based upon this relationship, the toxicity of sampling water could be distinguished. In this work, different working voltages were applied to improve the identification of wastewater’s toxicity in accordance with the output voltage. The characteristic voltage-time curves were fairly interpreted using three types of calculation methods, with which specific inhibition rates (I%) were thereafter expressed. Using the calculated I% close to the value measured by Oxygen Dissolved Analyzer (ODA) and the shortest response time of such method, an optimized microbial sensor with such built-in processing algorithm was provided. With the optimized reactivation time and bioactivity for the bacteria attached on the electrode, the analytical result demonstrated that the optimized working voltages, which provoked the maximum oxidation-reduction reaction rate or oxygen uptake rate of bacteria, for different bacteria-attached electrodes were 0.85 V for Bacillus subtilis, 0.65 V and 0.70 V for Escherichia coli, 0.70 V and 0.75 V for Pseudomonas putida, respectively. Taking phenyl solutions as the reference, these three bacteria were capable to detect the solutions in the ranges of 400-320 mgl-1 for Bacillus subtilis, 200-1600 mgl-1 Escherichia coli, 400-2400 mgl-1 Pseudomonas putida, respectively. In addition, the calculation models exhibited significant variations in determining phenyl’s toxicity with respect to the result of ODA. Using the best-fitted calculation model, the correlation coefficients were 0.951 for Bacillus subtilis, 0.903 for Escherichia coli, 0.921 for Pseudomonas putida, respectively.