大量傳統或是新興汙染物釋放至環境當中，可能對環境中的生物造成危害，因此需要藉由大量的毒理及生態毒理試驗來評估化學品的安全性，然而使用傳統毒理試驗會面臨許多挑戰，最主要的困境是使用大量實驗動物、耗費時間、金錢及嚴重缺乏生態毒理的數據，針對環境毒理試驗中的魚類急毒性而言，為達到避免使用動物與減少成本的目的，使用分層測試策略來評估魚類急毒性可能是一個具有高度測試效率的方式，在過去的毒理試驗研究當中，分層測試策略採取優先以電腦預測模式評估物質毒性，接著依序使用體外試驗，最後進行動物試驗，因此本研究目的旨在建立一個評估魚類急毒性的次序測試策略 (STS) ，並探索此策略在魚類急毒性上的預測力。研究將分為兩部分，第一部分為建立次序測試策略 (STS) ，收集文獻數據驗證整體策略之預測效力，第二部分為測試策略的應用，篩選缺乏替代試驗數據之物質，實際操作替代試驗得到測試策略預測之毒性結果，而電腦預測使用QSAR Toolbox 4.4.1、細胞株試驗採用OECD TG249、胚胎試驗採OECD TG236及魚類試驗採OECD TG203，並根據GHS規範辨識物質對魚類急毒性之危害。根據分層測試策略建立了魚類急毒性的次序測試策略估算物質毒性，使用文獻驗證物質應用於 STS 之危害辨識準確率為 84.62 %，其中61.54 % 物質採用替代試驗之數據；採用未知毒性物質實際進行STS準確率為 70.59%，61.54 % 物質沒有採用到體內試驗估計毒性。本研究藉由分層測試策略的概念建立魚類急毒性試驗的次序測試策略評估物質毒性，我們驗證 STS 之危害辨識準確率高達 84.62%，並且應用次序測試策略的同時可大幅度降低的動物使用量，綜合以上本研究提供一種魚類急毒性試驗的次序測試策略，這將對於生態毒理學的發展上有進一步改善單一替代試驗預測力不足之貢獻。

The practice of identifying the hazards of substances through alternative tests can alleviate the ethical controversies of traditional animal toxicity tests and shorten the experimental process. For the category of aquatic toxicity of acute fish toxicity, the single embryo test can not completely replace the animal test, and need to be combined with other methods. In this study, we aim to establish a Sequential Testing Strategy (STS) for acute fish toxicity, and explore the predictive efficacy. We combine the concept of stratified testing strategy, and estimate the toxicity of substances from fish-related in vitro to in vivo test. Moreover, we systematically determine the effects of aquatic toxicity Mode of Action on STS. The hazard identification accuracy rate of the STS for acute fish toxicity were as high as 70.59%. Furthermore, STS greatly reduces the number of applicative animals. However, we found STS by specific Mode of Action cannot significantly enhances the predictive effectiveness. In summary, this study provided a novel Sequential Testing Strategy for acute fish toxicity tests, which would contribute to further improving the predictive power of single acute fish alternative tests, and enhances animal welfare in the development of ecotoxicology.

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