| 摘要: | 烷基苯酚聚氧乙基醇(alkylphenol polyethoxlates, APEOn )為一廣泛使用於工業、農業與家用活動之非離子性界面活性劑，而辛基苯酚(octylphenol)與壬基苯酚(nonylphenol)為這類非離子性界面活性劑在自然界代謝後之主要產物，會擾亂內分泌系統之荷爾蒙分泌、合成、代謝與受體結合，對生物體之成長、發育及生殖等產生不良影響，此類化學物質統稱內分泌干擾物質（Endocrine Disruptors, EDs），俗稱環境荷爾蒙。辛基苯酚與雌激素受體結合能力為烷基苯酚聚氧乙基醇代謝物中之最高者，且誘導魚類產生卵黃前質能力為雌激素之4×10-4倍，雌激素效應是壬基苯酚的十至二十倍，因此本研究以octylphenol為篩選目標。本實驗室先前已篩選出17株能以辛基苯酚為唯一生長碳源之菌株，其中Methylobacterium radiotolerans SH771由含有辛基苯酚的藥罐分離出來，並以此菌株做後續的研究。本研究的目的是利用高效能液相層析質譜儀偵測Methylobacterium radiotolerans SH771的代謝物以探討其代謝途徑。液相層析圖譜中可發現唯一的碳源辛基苯酚濃度隨著培養時間增加而逐漸下降，且發現另一化合物1,2,4-trihydroxybenzene的形成。本研究推測辛基苯酚可能經由兩條路徑分解，其一為中央斷裂成2,4,4-trimethyl-2-pentanol和hydroquinone，另一途徑是經由苯環加氧酵素(Aromatic ring-hydroxylating oxygenase)產生中間代謝物 octylcatechol，再經中央斷裂，使烷基鏈與catechol相接處分開，繼而產生1,2,4-trihydroxybenzene及2,4,4-trimethyl-2-pentanol。Hydroquinone也會代謝成1,2,4-trihydroxybenzene。1,2,4-trihydroxybenzene之後再經由苯環切割酵素（Aromatic ring cleavage oxygenases）將其代謝物開環。中央斷裂和開環都會使其環境荷爾蒙效應去除，達成生物降解途徑之效果。 Alkylphenol polyethoxylates (APEOn) are an important group of non-ionic surfactants used in the agricultural, industrial and domestic detergents, and many other applications. In addition, nonylphenol and octylphenol are known as the major metabolites from APEOn. The prior studies have demonstrated that octylphenol is 10 to 20 folds higher in the estrogenic effect than nonylphenol. 4-t-octylphenol was used as the sole carbon source for the isolation of octylphenol-degrading bacteria in Taiwan. Seventeen octylphenol-degrading bacteria were isolated from paddy farm topsoil, soil microcosm and 4-t-octylphenol reagent bottles by previous works. Methylobacterium radiotolerans SH771 is belong to alpha proteobacteria and was isolated from a reagent bottle containing 4-tert-octylphenol in our lab. In many reports had shown Methylobacterium radiotolerans has biotechnology and bioremediation potential. The aim of this study was to analyze the key metabolites from octylphenol degradation by Methylobacterium radiotolerans SH771 and propose the degradation pathway. Octylphenol and its metabolites were analyzed by high-performance chromatography (HPLC) system equipped with an electrospray ionization-mass spectrometer (MS). The chromatogram was shown the octylphenol gradually decreased with time and produced 1,2,4-trihydroxyenzene. We proposed that the octylphenol can go two pathways then transformed to the ring cleavage compound. The first octylphenol would central cleavage into 2,4,4-trimethyl-2-pentanol and hydroquinone. The other one, octylphenol can via the aromatic ring-hydroxylating oxygenase, multicomponent phenol hydroxylase (Tuan et al., 2011), transformed octylphenol to octylcatechol (OC) then central cleavage to 1,2,4-trihydroxybenzene and 2,4,4-trimethyl-2-pentanol. 1,2,4-trihydroxybenzene is a common metabolite of this two pathway then ortho-cleavage into maleylacetate and meta-cleavage into semialdehyde. The estrogenic activity of octylphenol is decreased due to the central cleavage and ring fission by aromatic ring cleavage dioxygenases because of the collapse of the phenolic and hydrophobic moiety that interact with the estrogen receptor. However, the mechanism is remaining unclear. The octylphenol degradation pathway is anticipated to be completed in Methylobacterium radiotolerans. |
