Dissipation of sulfonylurea herbicides chlorsulfuron and imazosulfuron in soil and the effect on soil bacterial community structure

本研究探討兩種硫醯尿素類除草劑Chlorsulfuron及Imazosulfuron於土壤中之消散，並結合核酸萃取、聚合酶鏈鎖反應 (polymerase chain reaction) 及變性梯度凝膠電泳 (denaturaing gradient gel electrophoresis, DGGE) 等分子生物技術來探討兩種硫醯尿素類對於土壤中細菌族群結構之影響。試驗土壤採集自桃園區農業改良場 (簡稱Pu土壤) 及花蓮區農業改良場 (簡稱Wl土壤)，於土壤中添加濃度為50 mg kg-1 及10 mg kg-1 的除草劑，於30℃下避光孵育，定期萃取並以高效能液相層析儀分析土壤中除草劑殘量；同時以市售土壤核酸物質萃取試劑萃取土壤DNA，藉由聚合酶鏈鎖反應擴增其中16S rDNA的基因片段，以變性梯度凝膠電泳觀察菌相結構之變化。果發現化學水解及生物性降解為於環境中的主要降解途徑，其中除草劑Chlorsulfuron的半衰期介於6.8至28.4天，除草劑Imazosulfuron之半生期介於6.4至14.6天，而在相同條件處裡下在兩種除草劑在微酸性的Pu土壤中 (pH 5.24) 的半生期皆小於在偏中性的Wl土壤 (pH 7.52) 中的半生期。由PCR-DGGE指紋圖譜發現，部份於Pu土壤中濃度為10 mg kg-1 的除草劑Chlorsulfuron處理下圖譜上的亮帶，於濃度為50 mg kg-1 的處理中並未能觀察到或不明顯，表示高濃度處理的確會抑制其中特定微生物之生長；而由聚類分析發現，在Pu土壤與Wl土壤中，兩種除草劑在濃度為50 mg kg-1 的處理中，不同時間之間的菌相結構歧異度大，表示高濃度處理對於不同時間的菌相結構影響較大。In order to investigate the effect of soil bacterial community structure of two sulfonylurea herbicides including chlorsulfuron and imazosulfuron. Extraction of total genomic DNA, polymerase chain reaction (PCR), and denaturing gel electrophoresis (DGGE) are composed of this study to analyze the relationship between bacterial community during 70 days. The soils used in this study are collected from Taoyuan District Agricultural Research and Extension Center (soil Pu) and Hualien District Agricultural Research and Extension Center (soil Wl). The main different property of these two soils is pH, that soil Pu is a mild acidic soil (pH 5.24) and soil Wl is nearly neutral (pH 7.52). Applied with 10 and 50 mg kg-1 of each herbicide respectively and incubated at 30℃ in the dark. The treated soils were extraction by acetonitrile and analysed by HPLC-UV detector.esults showed that chemical hydrolysis and biodegradation are both the main pathway of these two herbicides. The degradation profile fits well with first-order dynamic equation, that the half-life values of chlorsulfuron are between 6.8 to 28.4 days and imazosulfuron with 6.4 to 14.6 days. Both of these two herbicides have a higher degradation rate in soil Pu, it seems because of an acidic-catalyzed degradation. The DGGE fingerprints shows that bacterial community structure has a higher diversity in the treatment of 50 mg kg-1 and the high replication rate indeed reduce some bacterial in the soil.中文摘要..........................................I文摘要.........................................II錄............................................III目錄...........................................IV目錄...........................................VI、前言..........................................1、文獻回顧......................................2一) 農藥與環境...................................2二) 硫醯尿素類除草劑簡介.........................5三) 本研究相關生物技術簡介.......................9、研究目的.....................................16、材料與方法...................................17一) 硫醯尿素類除草劑於土壤中的降解試驗..........17二) 硫醯尿素類除草劑對於土壤細菌族群影響之分析..27、結果與討論...................................33一) 硫醯尿素類除草劑於土壤中的降解試驗..........33二) 硫醯尿素類除草劑對於土壤細菌族群影響之分析..42、結論.........................................56考文獻.........................................5