特定益生菌降解食品中拟除虫菊酯类农药中间产物—3-苯氧基苯甲酸的特性及其机理研究

Pyrethroid pesticides have been widely applied in agricultural production. But their residues in foods are still unresolved. This not only threatens to human being's health but also restricts the sustainable development of modern agriculture and food industry. Based on current research status and our preliminary work on the selection of ideal probiotics with strong degradation activity, our research will focus on the further biodegradation process of their hydrolysis intermediate product, 3-phenoxybenzoic acid, so as to find out the key enzymes and the regularity of changes in its chemical structure and to reveal the pathway and mechanism of its biodegradation. Meanwhile, based on the degradation characteristics of different strains, we will study the influence of chemical composition in food and synergistic actions between the probiotics or the enzymes on the biodegradation of pyrethroid pesticides in order to establish a model for degrading residual pyrethroid pesticides in foods efficiently by the probiotics and their enzymes. The results will not only lay a foundation for estabishing the system of reducing and eliminating pyrethroid residues in foodstuffs by probiotics, but also provide valuable references to the biodegradation of other kinds of pesticides which can result in food unsafety. Therefore, the research possesses significant theory and practice values.

拟除虫菊酯类农药是目前广泛使用的杀虫剂之一，但消除其食品中残留的问题尚未解决，严重威胁着人类的身体健康，制约现代农业和食品工业的可持续发展。基于国内外研究现状和本课题组的前期工作（已筛选获得对拟除虫菊酯类农药具有较强降解活性的理想益生菌株）基础上，重点研究特定益生菌对拟除虫菊酯类农药的降解中间产物-3-苯氧基苯甲酸的生物降解特性和过程，确定其分子结构的变化规律和参与反应的关键酶，从而揭示其生物降解的途径和机理。同时，基于不同益生菌的降解特性，阐明复合益生菌或其酶的协同作用和食品组分对其降解效果的影响，确定益生菌及其酶制剂高效降解食品中拟除虫菊酯类残留农药的模式。课题研究成果不仅可为益生菌减少或消除食品原料中拟除虫菊酯类残留农药降解体系的建立奠定理论基础，而且也可为食品安全中亟待解决的其它残留农药和兽药的生物降解提供借鉴，具有较大理论和现实意义。

拟除虫菊酯类农药广泛使用带来的农产品中残留问题尚未得到解决，制约了现代农业和食品工业的可持续发展。3-苯氧基苯甲酸是多数拟除虫菊酯类农药（β-氯氰菊酯等）降解中间产物之一，对人和动物具有一定毒性，自然条件下难降解，半衰期长达180 d。本项目对源自不同生境微生物降解β-氯氰菊酯和3-苯氧基苯甲酸的特性进行了分析，重点研究了发酵食品或饲料中常用的乳酸菌、米曲霉、地衣芽孢杆菌等降解3-苯氧基苯甲酸的特性、途径、机理和菌株共培养、共代谢、基质调控的效果。结果显示，地衣芽孢杆菌B-1对β-氯氰菊酯降解能力较强，但不降解3-苯氧基苯甲酸；米曲霉M-4对3-苯氧基苯甲酸降解能力强，但不降解β-氯氰菊酯；黑曲霉-YAT等对二者均具有较好的降解效果；乳酸菌对二者基本无降解作用。米曲霉M-4降解3-苯甲氧基苯甲酸的酶属于诱导酶，产生的木质素过氧化物酶催化其羟基化生成3-羟基-5-苯氧基苯甲酸，细胞色素P450和木质素过氧化物酶的作用使该中间产物醚键断裂生成苯酚和3,4,5-三羟基苯甲酸，苯酚在木质素过氧化物酶作用下羟基化生成邻苯二酚，邻苯二酚和3,4,5-三羟基苯基酸在虫漆酶等作用下开环生成烯烃类化合物。地衣芽孢杆菌B-1与米曲霉M-4共培养使β-氯氰菊酯半衰期缩短了62.66%，3-苯氧基苯甲酸产生系数从0.846减小到0.001；地衣芽孢杆菌B-1和鞘氨醇单胞菌SC-1共培养，对较低浓度β-氯氰菊酯几乎完全降解，体系中3-苯氧基苯甲酸含量减少了55.5%。基质调控是提高菌株共代谢降解3-苯氧基苯甲酸效率的有效途径之一，分别添加葡萄糖、蛋白胨、尿素、硫酸铵、Fe3+、Fe2+或4-香豆酸、藜芦醇、高碘酸钠等可使米曲霉M-4共代谢对其降解效率有不同程度提高；表面活性剂—聚氧乙烯月桂醚可增加降解体系液相中β-氯氰菊酯含量，使地衣芽孢杆菌B-1对其降解率提高46.95%，半衰期缩短70%。基因重组是构建可降解该类农药的植物乳杆菌可能手段之一。课题研究结果有助于全面认识微生物降解3-苯氧基苯甲酸的途径和机理，为降解农产品或食品中残留农药的实际应用提供指导。