辛基苯酚聚氧乙基醇分解菌之多樣性及Pseudomonas sp. TX1代謝物之分析;The diversity of octylphenol polyethoxylate-degrading bacteria and the metabolites from Pseudomonas sp. TX1

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題名:	辛基苯酚聚氧乙基醇分解菌之多樣性及Pseudomonas sp. TX1代謝物之分析;The diversity of octylphenol polyethoxylate-degrading bacteria and the metabolites from Pseudomonas sp. TX1
作者:	林怡雯;Yi-Wen Lin
貢獻者:	生命科學研究所
關鍵詞:	辛基苯酚聚氧乙基醇;生物降解;環境荷爾蒙;Pseudomonas sp. TX1;octylphenol polyethoxylate;biodegradation;environmental hormone;Pseudomonas sp. TX1
日期:	2009-07-22
上傳時間:	2009-09-22 10:21:13 (UTC+8)
出版者:	國立中央大學圖書館
摘要:	辛基苯酚聚氧乙基醇 (Octylphenol polyethoxylates, OPEOn)為一聚氧乙烯鏈鏈長從4到15，平均鏈長9.5的混合物，商品名為Triton X-100，屬於非離子性界面活性劑的一種，被廣泛使用於工業應用、農業及家庭清潔劑中。此研究中從農藥工廠、稻田及中央大學排放水的表土及底泥中分離出43株菌株，以95種不同碳源組合圖譜（BioLog方法）及16S rRNA序列比對作菌種鑑定。其中除了Pseudomonas這屬的菌株外，Moraxella osloensis、Aquaspirillum peregrinum ss integrum、Brevibacterium iodinum、Photobacterium logei、Ochrobactrum sp.、Cupriavidus sp.、Ensifer adhaerens、Achromobacter xylosoxidans及Brevbacterium iodinum皆尚未被發表過具有分解OPEOn的能力；其中，B. iodinum為唯一的一株革蘭氏陽性菌。農藥工廠溫室中分離中的菌相複雜度大於稻田，而稻田的菌相複雜度又大於中央大學排放水底泥。初步菌株活性分析，以OPEOn為唯一碳源培養之菌體測定對OPEOn、同樣具有聚氧乙烯鏈的dodecyl octaethoxylate (AEO8)及其推測可能之代謝產物之消耗氧氣的能力。依菌株對於不同化合物具有不同的耗氧活性，將其分成三類：第一類菌株對於OPEOn及AEO8具有較高耗氧活性，其中包括Pseudomonas sp. TX1；第二類菌株則對OPEOn具較高耗氧活性，但對AEO8則無；第三類菌株只有B. iodinum，為唯一對具雌激素活性化合物辛基苯酚(octylphenol, OP)具耗氧活性的菌株。在43株具分解OPEOn能力的菌株中，Pseudomonas sp. TX1生長素率最快且對OPEOn的耗氧活性最高，並能生長在0.05%到20%的OPEOn濃度中，其生長速率在0.34到0.44 hr-1之間。以高效能液相層析質譜儀(HPLC-MS)分析菌株TX1分解OPEOn的代謝產物，推斷此菌株能減短聚氧乙烯鏈並生成OP。在此菌株生長的對數期(log phase)有短鏈的羧酸化產物生成，例如在烷基鏈上產生羧酸化的carboxyoctylphenol polyethoxylates (COPEOn, n = 2, 3)及雙羧酸化產物carboxyoctylphenol polyethoxycarboxylates (COPECn, n = 2, 3)；在聚氧乙烯鏈產生羧酸化產物octylphenol polyethoxycarboxylates (OPECn, n = 1-3)則在菌株生長進入靜止期(stationary phase)後生成。所有短鏈的代謝物：OPEOn、OPECn、COPEOn及COPECn皆在靜止期有累積的現象。在文獻中尚未有研究發表過由好氧菌分解OPEOn而生成在烷基鏈上產生羧酸化產物COPEOn及雙羧酸化產物COPECn的現象，Pseudomonas sp. TX1為第一株被發現據此能力的菌株。 Octylphenol polyethoxylates (OPEOn), also known as Triton X-100, a nonionic surfactant, was widely used in industrial, agricultural, and domestic applications. It is a mixture composed of OPEOn (n = 4~15) with an average unit of ethoxylate at 9.5. In this study, forty three bacterial strains were isolated from soil samples and sediments of pesticide factory, rice field, drainage of dormitory on campus. They were identified by two methods, Biolog breathprinting and 16S rDNA sequence analysis. Except species from Pseudomonas, other bacteria included Moraxella osloensis, Aquaspirillum peregrinum ss integrum, Brevibacterium iodinum, Photobacterium logei, Ochrobactrum sp., Cupriavidus sp., Ensifer adhaerens, Achromobacter xylosoxidans, and Brevbacterium iodinum have not been reported to be capable in the degradation of OPEOn. B. iodinum was the only Gram-positive bacterium. The diversity of the isolated strains from the green house of a pesticide factory is higher than the rice field and the drainage of NCU campus. In this study, we apply the oxygen consumption assay to evaluate the aerobic degradation of OPEOn. Depending on the oxygen consumption activity of the cells incubated with different substrates, the isolates are separated into three groups. Group I: the oxygen consumption activities on OPEOn and AEO8 are high, including Pseudomonas sp. TX1. Group II: the activity is high on OPEOn, but low on AEO8. Group III contains only one Gram-positive strain, B. iodinum, is the only strain in our isolates showed oxygen consumption activity toward ocylphenol, which is a recalcitrant compound and is normally detected in a variety of environments. Among the isolates, Pseudomonas sp. TX1 can grow on 0.05% to 20% of OPEOn with a specific growth rate of 0.34-0.44 hr-1. High-performance liquid chromatography–mass spectrometer analysis of OPEOn degraded metabolites revealed that strain TX1 was able to shorten the ethoxylate chain and produce octylphenol (OP). Furthermore, formation of the short carboxylate metabolites, such as carboxyoctylphenol polyethoxylates (COPEOn, n = 2, 3) and carboxyoctylphenol polyethoxycarboxylates (COPECn, n = 2, 3) began at the log stage, while octylphenol polyethoxycarboxylates (OPECn, n = 1-3) was formed at the stationary phase. All the short-ethoxylated metabolites, OPEOn, OPECn, COPEOn, and COPECn, accumulated when the cells were in the stationary phase. This study is the first report to demonstrate the formation of COPEOn and COPECn from OPEOn by an aerobic bacterium.
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