噻嗪酮降解菌株的筛

发布时间：2018-03-22 00:19

本文选题：噻嗪酮　切入点：降解　出处：《淮北师范大学》2017年硕士论文　论文类型：学位论文

【摘要】：噻嗪酮是一种新型的杀虫剂,能够通过抑制几丁质的合成干扰昆虫的新陈代谢,主要用于对飞虱、粉虱、叶婵及介壳虫等同翅目害虫有很好的防治作用。由于噻嗪酮在使用过程中的不规范储藏,和在农作物上的重复喷洒,对环境造成了危害。因此,如何治理环境中的噻嗪酮残留就成为了我们急需解决的难题。可通过物理方法、化学方法、生物方法治理环境中的噻嗪酮污染,生物降解是治理的重要方法之一,高效降解菌能够将噻嗪酮作为碳氮源将其代谢成小分子物质,在生物修复中起着重要的作用。遗憾的是,有关噻嗪酮降解菌的报道只有五株,有关其降解途径的报道也少之又少。本研究从常年生产噻嗪酮农药的工厂排污口处采取水样与土样,于100 m L无机盐培养基中,并添加噻嗪酮作为碳源,在30°C、160 rpm min-1摇床条件下经富集驯化和分离纯化,成功筛选到多个高效降解菌。对具有噻嗪酮降解能力的菌株做了降解能力的验证,选择一株降解能力较好的菌株展开研究,将其命名为RX-3。通过形态学、生理生化实验及16S rRNA序列比对鉴定菌株RX-3为红球菌属(Rhodococcus sp.),其最适生长温度为30°C,最适生长酸碱度值为7.0,且对金属离子Ba2+、Zn2+和Cu2+有一定的耐受性。当无机盐中噻嗪酮浓度为60 mg L-1时,菌株RX-3在80 h内能够降解98.01%的噻嗪酮。使用二氯甲烷等体积萃取法提取到噻嗪酮的降解产物,并借助于气相色谱质谱联用仪分析了9种中间代谢产物,初步剖析了噻嗪酮的降解途径。在代谢途径A中,噻嗪酮中的杂环发生水解生成中间代谢产物B;在代谢途径B中,噻嗪酮的代谢起始于侧链中叔丁基和异丙基键的断裂,伴随产物C,E和F的生成,随后发生了水解反应和氧化还原反应。在代谢途径C中,噻嗪酮的代谢起始于苯环与N之间化学键的断裂,生成中间代谢产物D。本研究使用聚乙二醇4000对噻嗪酮降解菌和毒死蜱降解菌的原生质体进行融合,成功获得其融合子。
[Abstract]:Thiazinone is a new type of insecticide that interferes with the metabolism of insects by inhibiting the synthesis of chitin. It is mainly used in the treatment of planthopper and whitefly. Homoptera pests such as Ye Chan and mesoptera insects have a good control effect. Because of the nonstandard storage of thiazinone in the process of use and repeated spraying on crops, it is harmful to the environment. How to control thiazinone residues in the environment has become a difficult problem that we need to solve urgently. The biodegradation is one of the most important methods to control thiazinone pollution in the environment by physical, chemical and biological methods. Highly efficient biodegradable bacteria can metabolize thiazines as carbon and nitrogen sources into small molecules and play an important role in bioremediation. Unfortunately, only five strains of thiazone degrading bacteria have been reported. In this study, water samples and soil samples were taken from the sewage outlets of factories that produce thiazinone pesticides all the year round, and thiazinone was added as carbon source in 100 mL inorganic salt medium. After enrichment, acclimation, purification and purification, a number of high efficient biodegradable bacteria were successfully screened under the condition of 30 掳CX 160 rpm min-1 shaking bed. The biodegradable ability of the strain with thiazinone degradation ability was verified, and a strain with better degradation ability was selected. It's named RX-3. it's morphologically, Physiological and biochemical experiments and 16s rRNA sequence alignment identified the strain RX-3 as Rhodococcus sp., the optimum growth temperature is 30 掳C, the optimum growth pH value is 7.0, and the strain is tolerant to metal ions Ba2 Zn 2 and Cu2. When the concentration is 60 mg L ~ (-1), The strain RX-3 could degrade 98.01% thiazinone within 80 h. The degradation products of thiazinone were extracted by dichloromethane isovolumetric extraction, and 9 intermediate metabolites were analyzed by gas chromatography-mass spectrometry. The degradation pathway of thiazinone was preliminarily analyzed. In metabolic pathway A, the heterocycles in thiazinone hydrolyzed to form intermediate metabolite B, and in metabolic pathway B, the metabolism of thiazinone originated from the breakage of tertiary Ding Ji and isopropyl bond in the side chain. With the formation of Con E and F, hydrolysis reaction and redox reaction took place. In metabolic pathway C, the metabolism of thiazinone begins with the breaking of the chemical bond between benzene ring and N. In this study, the protoplasts of thiazine-degrading bacteria and chlorpyrifos degrading bacteria were fused with polyethylene glycol 4000, and the fusion was successfully obtained.
【学位授予单位】：淮北师范大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X172;X592

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