【作者】 邹月利；

【导师】 陶波；

【作者基本信息】 东北农业大学 ， 农药学， 2012， 硕士

【摘要】 氯嘧磺隆(Chlorimuron-ethyl)[2-(4-氯-6-甲氧基嘧啶-2-基氨基甲酰氨基磺酰基)苯甲酸(乙酯)]是美国杜邦公司于20世纪80年代开发的一种磺酰脲类除草剂。氯嘧磺隆是一种超高效、广谱的大豆田苗后除草剂,对一年生禾本科杂草、部分阔叶杂草及部分莎草科杂草具有较好的防治效果。氯嘧磺隆在土壤中残留时间长,并且极低残留量就可以引起后茬敏感作物的药害,严重影响农业生产中种植结构的调整。微生物能够明显降解除草剂残留,并且对环境安全、无毒。因此,利用微生物治理此类长残留除草剂的污染,在工农业生产领域中具有非常重要的意义。本文利用仪器分析和生物化学方法系统的研究了,在不同条件下真菌对氯嘧磺隆除草剂的降解作用;采用HPLC结合LC-MS对氯嘧磺隆的主要降解产物进行初步鉴定,明确了氯嘧磺隆除草剂在不同条件下的降解机理和代谢途径。主要研究结果如下：1.黑曲霉在基础培养基中最适宜的培养条件：氯嘧磺隆浓度为10.0mg·L-1,温度为30.0℃,接种量为5.0mL。2.不同真菌菌株对氯嘧磺隆的降解速率不同。顺序为：黑曲霉>黄曲霉>F8。其中,真菌黑曲霉对氯嘧磺隆的降解率为96.49%,黄曲霉为88.95%,F8为82.37%。3.混合菌株对氯嘧磺隆的降解速率均高于其单一菌株。顺序为：黑曲霉与黄曲霉混合菌株>黑曲霉与F8混合菌株。两种混合菌株对氯嘧磺隆的降解率均在98.2%以上。4.在不同真菌菌株的作用下,随着氯嘧磺隆降解率的增加,降解液的pH值逐渐降低。其中,加入黑曲霉的氯嘧磺隆降解液pH值由最初的7.55降低到3.42；加入黑曲霉与黄曲霉混合菌株的降解液pH值变化幅度最大,由最初的7.51降低到3.27。5.氯嘧磺隆的水解和微生物降解产物不同。其中,水解和微生物降解产物均含有产物A[N,N-二邻甲酸乙酯苯磺酰基尿素]、B[邻甲酸乙酯苯磺酰胺基甲酸酯]、C[邻磺酰胺苯甲酸乙酯]、D[邻磺酰苯酰亚胺]、G[2-(4-氯-6-甲氧基嘧啶-2-基氨基甲酰氨基磺酰基)苯甲酸]和J[2-(4-氯-6-甲氧基嘧啶-2-基亚胺醇氨基磺酰基)苯甲酸乙酯]。此外,真菌黑曲霉对氯嘧磺隆的降解产物还有产物E[2-(氨基磺酰基)苯甲酸]、F[邻甲酸乙酯苯磺酰基异氰酸酯]和H[2-氨基-4-氯-6-甲氧基嘧啶];真菌黄曲霉对氯嘧磺隆的降解产物还有产物F；真菌黑曲霉与黄曲霉混合菌株对氯嘧磺隆的降解产物还有产物F和H。6.氯嘧磺隆的水解和微生物降解机理及代谢途径不同。虽然水解和微生物降解反应都是苯环上的酯基水解和磺酰脲桥断裂,转化为2-氨基-4-氯-6-甲氧基嘧啶和2-(氨基磺酰基)苯甲酸乙酯等物质；甲氧基嘧啶环可开环被降解为CO2和H20；嘧啶环上甲氧基稳定,没有脱甲氧基反应发生。但是微生物对氯嘧磺隆的降解机理更为复杂,真菌菌株不仅可以通过分解水解的产物来促进水解反应的进行,而且可以使生成的裂解产物进一步发生水解、氧化、醇解和环化反应；其中,黑曲霉、黑曲霉与黄曲霉混合菌株还可以分解氯嘧磺隆的互变异构体,使分解产率更高。更多还原

【Abstract】 Chlorimuron-ethyl, ethyl2-(4-chloro-6-methoxypyrimidin-2-ylcarbamoylsulfamoyl) benzoate, is a sulfonylurea herbicide discovered and developed by Du Pont campany of United States in1980s. Chlorimuron-ethyl, as a super-effective and broad-spectrum postemergence herbicide controlling soybean field weeds, had good effect on the control of annual grass weeds, some broadleaf weeds and some sedge weeds. But chlorimuron-ethyl degraded very slowly and remained for long time in soil, which could cause injury to succeeding sensitive crop. Microorganisms can significantly degrade herbicide residues without environmental problem and toxicity. Therefore, the use of microbial treatment has a very important theoretical significance and great potential to chlorimuron-ethyl residual pollution. A solid phase extraction-high performance liquid chromatography-mass spectrometry method and biochemical assay method were established for determination the degradation of chlorimuron-ethyl herbicide by fungi strains at different conditions in this paper. Main degradation products of chlorimuron-ethyl were studied with HPLC, and LC-MS respectively. The mechanism and pathway of degradation were studied under different conditions.The results showed that:1. Optimum conditions for degradation of chlorimuron-ethyl herbicide by Aspergillus niger were as follows:concentration of chlorimuron-ethyl in basal medium was10mg·L-1, temperature of reaction solution was30℃, inoculum of Aspergillus niger was5.0mL.2. Degradation rates of different fungi strains were very differently. The order was:Aspergillus niger>Aspergillus flavus>F8. The results showed that the highest degradation yield was produced by Aspergillus niger (96.49%), followed by Aspergillus flavus (88.95%), while F8was the lowest (82.37%).3. Degradation rates of mixed fungi strains were higher than single fungi strain. The order was: Aspergillus niger and Aspergillus flavus>Aspergillus niger and F8. Degradation yields of these mixed fungi strains were higher than98.2%.4. Using different fungi strains, the pH of reaction solution reduced in pace with degradation of chlorimuron-ethyl. The pH value of Aspergillus niger reaction solution reduced from the initial7.55to3.42and the pH value of Aspergillus niger and Aspergillus flavus reaction solution reduced enormously from the initial7.51to3.27.5. Degradation products of hydrolysis and biodegradeation were different. Product A、B、C、 D、G、J were contained commonly in hydrolysis and biodegradeation. In addition, product E、F and H were contained in degradation by Aspergillus niger; product F was contained in degradation by Aspergillus flavus; product F and H were contained in degradation by Aspergillus niger and A spergillus flavus.6. The degradation mechanism and pathway of hydrolysis and biodegradeation were different. Chlorimuron-ethyl was degraded by means of hydrolyzed the ester group on the benzene ring and destroyed the sulfonylurea bridge of chlorimuron-ethyl. Methoxylpyrimidine ring was unstable and there was oxidation reaction occured. Pyrimidine ring was stable and there was no de-methoxy reaction occured. The mechanism of biodegradeation was more complex than hydrolysis. Hydrolysis products were degraded by fungi strains and degradation product could be further degraded by hydrolysis, oxidation, alcoholysis and cyclization reactions. At last, chlorimuron-ethyl isomer also degraded by Aspergillus niger, mixed fungi strains of Aspergillus niger and Aspergillus flavus, which made degradation yield higher.更多还原