以月芽藻類毒性試驗方法評估α,β-不飽和酮類之毒性與結構-活性關係之研究

Abstract

本篇研究以月芽藻 (Pseudokirchneriella subcapitata)針對18種酮類，其中16種為□□□-不飽和酮，2種為飽和酮，所進行之48小時密閉式毒性詴驗。實驗所得到之結果，將利用□Do、Final yield及Growth rate做為觀測終點，藉由Probit模式求出半致死濃度 (50% Effect concentration，EC50)，將其毒性試驗後所得到的log(1/ EC50)，與其物化參數(Log P)、電性參數(Delta E、Q3、Q4、Elumo、Ehomo)及反應性參數(log(1/ RC50))進行迴歸分析，找出定量-結構活性關係模式。 毒性由強至弱依序為不飽和酮＞鹵素取代酮＞飽和酮，炔酮類＞烯酮類，無取代基不飽和酮＞甲基取代酮，造成此趨勢之主因為酮類含C＝O，屬於拉電子基，造成本身電子雲密度不均，使得酮類本體不穩定；不飽和酮含□鍵，使酮類結構活躍，導致電子易發生轉移，與生物體內所含之硫氫基(－SH)產生交互作用，進而影響毒性；而甲基取代造成立體障蔽(steric hindrance)效應，使酮類不易與電子反應，毒性減弱。另外，由反應性試驗數據呈現亦與上述結果相同。 敏感度分析方面，藻類毒性試驗反應終點以最終細胞變化量最為敏感；與其他物種進行比較，藻類敏感性比纖毛蟲高。 QSAR分析發現，本詴驗毒物屬親電性物質，與麻醉性作用機制常選用之參數－辛醇-水係數迴歸結果不佳，故其不為反應性毒性作用機制；而利用反應性參數迴歸效果佳，毒性與反應性呈現正相關；由逐步迴歸分析法，挑選出與試驗毒物相關性佳之電性參數，也找到一些相關性高之方程式(R2=0.821~0.907)，未來可藉此預測迴歸式預測具相關結構、相同毒性作用機制之毒化物，以減少在生物毒性試驗上人力、時間、設備上之成本。

The objective of this study is to understand the toxic effect of ketones (that contains sixteen α,β-unsaturated ketones and two saturated ketones ) on Pseudokirchneriella subcapitata using a closed system test. The effects of ketones are evaluated by three kinds of response endpoints, i.e., final yield, growth rate, and the dissolved oxygen production. Median effective concentrations (EC50) are estimated using the Probit model with a test duration of 48hrs. Then, log (1/EC50) is utilized to make a regression with physical chemistry descriptors, electronic descriptors and reactive descriptors. Finally, we establish the QSAR models. The results show that unsaturated ketones are the toxicest of these ketones. Alkynones are more toxic than alkenones, and no substitution ketones are more toxic than Methyl substitution ketones. Ketones become unstable owing to electron-withdrawing effect caused by the existence of carboxyl of ketones. In addition, unsaturated ketones contain □-bond that make electron easily transfer and interact with sulfhydryl group which is in organisms. Methyl group substitution cause steric hindrance, hense toxicity decrease a lot. The same results of toxicity are also discovered in Reactivity. The order of the relative sensitivity is as follows：algae(Final yield)＞algae(Do production) ＞algae(Growth rate)＞Tetrahymena pyriformis. The toxicant of this study is electrophile which results in poor relationship with 1-octanol/water partition coefficient. Toxicity is positive correlation with reactivity. The QSAR models are established well (R2=0.821~0.907) that based on electronic descriptors by stepwise analysis. The results of this study can help us to decrease the cost of toxicity test.