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研究生:李昭緯
研究生(外文):Li, Jhao-Wei
論文名稱:應用電化學法於紅茶飲料中衛生指標微生物之檢測
論文名稱(外文):Quantitative Determination Of Microorganism In Black Tea Drinks Using Electrochemical Method
指導教授:徐詮亮徐詮亮引用關係
指導教授(外文):Hsu, Chuan Liang
口試委員:張谷昇詹鴻得
口試委員(外文):Chang, Ku ShangJang, Hung Der
口試日期:2012-06-08
學位類別:碩士
校院名稱:東海大學
系所名稱:食品科學系
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2012
畢業學年度:100
語文別:中文
論文頁數:78
中文關鍵詞:安培電化學法
外文關鍵詞:amperometric
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茶飲料是台灣最受歡迎的飲料之ㄧ,但常有生菌數過高、大腸桿菌群超標等問題,傳統檢測大腸桿菌及大腸桿菌群的標準方法包括:API system、Coliform agar及PetrifilmTM 等,但仍難擺脫繁瑣的步驟及耗時的過程。本研究主要以建立一套簡單快速、成本低廉的檢測方法為目的,實驗過程所使用的大腸桿菌群菌株包含Escherichia coli、Enterobacter、Citrobacter等,及Salmonella、Pseudomonas和Morganella三株非大腸桿菌群之革蘭氏陰性菌,操作條件分別設定在0.3及0.5V,並維持恆溫37℃,因為大腸桿菌培養過程所產生的代謝產物為電化學活性物質,達到一定濃度時可因通過電極之電流瞬間增加,產生明顯訊號,另外實驗中亦發現可藉由改變選擇性培養基成分達到分離大腸桿菌群及其他革蘭氏陰性菌的效果。結果顯示紅茶中之微生物濃度與檢測時間(Detection time, DT)呈線性關係,菌體濃度之檢測範圍可達101至108 CFU/ml ,且實驗中亦發現可藉由排除選擇性培養基中緩衝溶液之成份達到區分大腸桿菌及大腸桿菌群的效果。本研究開發之方法用於茶飲料總生菌數之檢測所需檢測時間根據實驗結果菌體濃度於102 CFU/ml以上均小於10小時,相較於傳統檢測方法需24至48小時,本方法明顯簡單、快速。



The contamination of highly plate counts including Escherichia coli in black tea drink, one of the most popular beverages in Taiwan, has been becoming a major concern for the public health. The traditional methods for detection of E. coli include API System, Coliform Agar and Pertrifilm, etc., however, these culturing methods are excessively time-consuming and take cumbersome steps. In this study, a simple, rapid and cost-effective analytical method for the detection of E. coli and coliforms was established. Cultures of E. coli, coliforms and non-colifoms gram-negative bacteria were tested using the electrochemical method. The amperometric measurements were performed in the selective medium by the addition of black tea drink at a working potential of 0.3 V or 0.5 V and temperatures at 37 ℃. The results showed that a linear relationship exists between the logarithm of the bacterial concentration (over the range of 101 to 108 CFU/mL) and the detection time (DT). The addition of black tea drink into the selective medium caused the release of electrochemical active substances in microorganisms and produced a significant signal through the electrode reaction. The correlation coefficient (R2) was 0.9354 and 0.9144 for the detection of E. coli and coliforms, respectively. The DT obtained from the proposed method is about 10 hours, which is shorter than those by traditional culturing methods. The proposed electrochemical method could both rapidly and sensitively determine the concentration of E. coli and coliforms in black tea drink samples.
目錄
頁碼
中文摘要...................................................................................................Ι
英文摘要................................................................................................... II
目錄.......................................................................................................... III
圖目錄..................................................................................................VI
表目錄..................................................................................................... VIII
第一章、前言. .........................................................................................1
第二章、文獻回顧.................................................................................. 3
2.1大腸桿菌與大腸桿菌群............................................................... 3
2.1.1特性描述.............................................................................. 3
2.1.2檢測方法..............................................................................4
2.2生物感測器...................................................................................7
2.2.1電化學生物感測器..............................................................7
2.2.2以電化學生物感測器開發多功能檢測平台之潛力..........10
2.2.3以電化學感測器檢測大腸桿菌之目前研究結果..............13
2.3紅茶...............................................................................................14
2.3.1紅茶中成分分析及保健功效..............................................14
2.3.2衛生署訂定飲料類法定標準..............................................17
第三章、材料與方法...............................................................................19
3.1儀器設備.......................................................................................19
3.2培養基...........................................................................................20
3.3化學試劑......................................................................................20
3.4茶飲料樣品...................................................................................21
3.5試驗用菌株...................................................................................21
3.6電化學檢測系統之架構...............................................................23
3.7循環伏安(Cyclic voltammetry, CV)掃描.....................................28
3.8菌株活化.......................................................................................29
第四章、結果與討論.............................................................................31
4.1 電化學檢測微生物機制之探討...................................................31
4.2電化學系統檢測微生物之最適電壓探討...................................36
4.3電化學系統檢測微生物之最適溫度探討...................................40
4.4以營養培養基檢測市售紅茶中微生物之探討...........................42
4.5以循環伏安法檢測市售紅茶中微生物之探討...........................44
4.6以電化學系統檢測市售紅茶中微生物之探討...........................46
4.7電化學系統檢測大腸桿菌及大腸桿菌群之培養基碳源及成分
探討...............................................................................................48
4.8電化學檢測大腸桿菌於不同糖度紅茶下生長之探討...............54
4.9電化學法及傳統方法對連鎖店紅茶飲料微生物之檢測...........56
4.9.1大腸桿菌與大腸桿菌群之檢測..........................................57
4.9.2總生菌數之檢測..................................................................58
第五章、結論.........................................................................................72
參考文獻.................................................................................................74


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