本研究利用假單胞菌(ATCC 23973)對化合物的瞬時耗氧速率來評估苯環單取代與酚類化合物的毒性，酚類化合物大都為非競爭型抑制劑，該類化合物的抑制常數Ki值代表的意義為抑制假單胞菌50% 瞬時耗氧速率的抑制劑濃度，因此，我們用Ki值做為化合物毒性的指標。
由於化學物質的毒性受到毒物進入生物相的穿透力以及與生物體之反應位置的相互作用的影響，因此，我們建立了Ki值與辛醇-水分配係數(logP)和最低未填滿分子軌域能量(LUMO)的結構-活性定量關係，並由此關係預測化合物對假單胞菌的抑制機制。
其中有一些化合物(pKa≦6.5)，它們的毒性偏低，因為該類化合物在實驗溶液中皆以共軛鹼(phenoxides)的形式存在，所以它們的毒性為共軛鹼所表現出的行為，而非如文獻上將其歸類的弱酸呼吸去偶合物，就是它們會抑制三磷酸腺苷的形成以及氧化鄰酸化的作用。
在與其他毒性測試方法的比較，發現本實驗結果與纖毛蟲的毒性測試有高度的相關性(r = 0.92)。

We have used the Pseudomonas putida (ATCC 23973) initial oxygen uptake (PIOU) data to assess the toxicity of the monosubstituted aromatic compounds and phenol derivatives. The inhibition constant, Ki, of monosubstituted aromatic compounds and phenol derivatives were measured and found that most phenol derivatives are belonging to the non-competitive inhibitors toward Pseudomonas putida. These Ki constant represent the concentration of the phenol derivatives, under which 50% of the oxygen uptake activity of Pseudomonas putida has been inhibited. Thus, Ki is used as an index to assess the toxicity.
The penetration ability of the toxic chemical into the organism and its following reaction site with the organism are use to assess the toxicity. Thus, the Ki value was correlated with both the n-octanol/water partition coefficient (log p) and the energy of lowest unoccupied molecular orbital (LUMO) to establish a quantitative structure-activity relationship (QSAR) model and found that this established equation allow one to predict the inhibition mechanism toward P.putida.
Several phenol derivatives ( pKa≤6.5 ) with lower toxicity previously called uncupler was also found not following the QSAR behavior, i.e. can not be explained using the established QSAR equation. The abnormal toxicity behavior of these compounds was found mainly due to having weak acid dissociation property into its conjugate base rather than due to being an uncoupler inhibiting the oxidative phosphorylation of ADP to form ATP.
Comparing our results obtained from PIOU method with that from other test methods, it was found that our results have a good relationship with that from a 2-day Tetrahymena pyriformis with r = 0.92.

中文摘要...................................i
英文摘要...................................ii
誌謝.......................................iii.
目錄.......................................v
表目錄.....................................x
圖目錄.....................................xi
第一章 緒論 ..............................1
一、生物分解...............................1
二、利用生物測試化合物的毒性...............2
(一)、魚類.................................3
(二)、MicrotoxTM...........................4
(三)、纖毛蟲(Tetrahymena pyriformis).......5
三、假單胞菌(Pseudomonas putida)的簡介.....5
四、酵素動力學(Enzyme kinetic).............6
(一)、Michaelis-Menten 方程式..............6
(二)、Briggs-Haldane穩定態處理法(steady-state treatment).................................8
(三)、Lineweaver-Burk方程式................9
(四)、酵素抑制之動力學模式.................10
1.競爭型抑制(competitive inhibition).......10
2.非競爭型抑制劑(noncompetitive inhibition)13
3.不競爭型抑制(uncompetitive inhibition)...14
五、假單胞菌瞬時耗氧速率測試...............16
六、結構-活性定量關係(Quantitative Structure-Activity Relationship；QSAR).....16
(一)、Hammett equation.....................17
1. 電子需求.............................17
2. Hammett equation.....................19
(二)、參數(parameters).....................21
1. 親油性參數(Hydrophobic parameters)...21
2. 電子的參數(Electronic parameters)....21
3. 空間的參數(Steric parameters)........22
(三)、QSAR的應用...........................22
七、毒性作用的模式和反應機構...............22
八、研究動機...............................24
第二章 實驗 ..............................25
一、藥品...................................25
(一) 培養細菌基質..........................25
(二) 抑制劑................................25
二、儀器...................................28
三、細菌培養...............................31
(一) 培養基的配置..........................32
(二) 微量元素溶液的配置....................32
(三) 轉菌..................................33
(四) 離心..................................33
四、氧電極的測試...........................33
(一) 溶液的配置............................34
1. 緩衝溶液的配製..........................34
2. 不同受質濃度的配製......................34
3. 不同受質濃度的配製......................34
(二)、瞬時耗氧速率的測試...................35
(三)、數據處理.............................36
五、分子參數與統計分析.....................38
第三章 結果 ..............................40
一、化合物的Ki值...........................40
二、競爭型抑制劑...........................41
三、結構-活性定量關係(Quantitative Structure-Activity Relationship；QSAR).....43
四、偏離點的分析...........................45
第四章 討論 ..............................47
一、假單胞菌瞬時耗氧速率方法(Pseudomonas Initial Oxygen Uptake，PIOU)的特色.................47
二、結構-活性定量關係(QSAR)................48
三、單取代化合物的討論.....................49
四、偏離點的分析...........................50
五、結構-活性定量關係的比較(comparative QSAR).52
(一)、假單胞菌與纖毛蟲(Tetrahymena pyriformis)毒性測試的比較.................................55
(二)、假單胞菌與鰷魚(Pimephales promelas)毒性測試的比較.......................................57
(三)、假單胞菌與Microtox毒性測試方法的比較.58
六、結論...................................59
參考文獻 ..................................60
附錄.......................................xiii
附表一、抑制劑濃度的配製
附表二、化合物PIOU實驗測試結果, Ki值以及物理-化學參數
附表三、抑制劑在各種毒性測試下之數據
PIOU實驗圖表

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