手性农药茚虫威对映体在茶叶生长加工过程中降解差异机制

In recent years, indoxacarb becomes the first choice for controlling leaf hoppers and other pests in the tea garden to replace other high toxic and high water-soluble insecticides. It has been reported that the S-enantiomer of indoxacarb has higher insecticidal activity and less toxicity than R-enantiomer, and the toxicity of metabolite DCJW was much higher than that of indoxacarb its own. During the study of our previous National Natural Science Foundation of China Youth Fund, we have found that the chiral pesticide-indoxacarb enantiomers selectively degraded in the growth and processing of tea. The S-enantiomer degraded faster than the R-enantiomer, that will lead to decreased efficacy and increased risk of residual toxicity, and the selective degradation mechanism is unclear.. Therefore, we propose to use metabolic profiling and omics strategies, with the help of UPCC/UPLC/APGC-QTOF-MS screening technology to study the differences of metabolites, migration and transformation for indoxacarb enantiomers in the growth, processing and brewing of tea, and the effects of different enzymes on the metabolism of indoxacarb enantiomers, so as to clarify the key metabolic enzymes, the key points and the mechanism of selective degradation differences for enantiomers at biochemical level..The project will provide scientific basis data for the promotion of chiral pesticides reasonable application in tea garden, and ensure the safety of tea drinking. It is also helpful for further reducing the application of chiral pesticide and improving the efficiency, reducing pesticide residue amounts in tea and improving the quality and safety of tea.

“十二五”以来，茚虫威逐渐成为防治茶园叶蝉等害虫、用于替代高毒、高水溶性农药的首选。据文献报道，茚虫威手性对映体中，S-体杀虫活性高且毒性比R-体小，而代谢物DCJW的毒性则大于茚虫威本身。在前面的青年基金研究中，我们发现了茚虫威在茶叶生长加工中存在对映体降解速率差异，S-体降解快于R-体,这样就会导致药效下降且残留毒害风险增大，对其中的选择性降解机制也不清楚。因此，本项目选择以茶叶为载体，利用代谢轮廓分析法和组学策略，借助UPCC/UPLC/APGC-QTOF-MS筛查技术，研究茚虫威对映体在茶叶生长-加工-冲泡中的代谢产物及迁移转化差异的规律、不同酶对对映体代谢的影响，从而找到关键控制点、明确关键代谢酶，从生化水平解析明确茚虫威对映体降解差异的机制。本项目为促进手性农药在茶园合理施用、保障茶叶饮用安全提供科学依据，有助于后续茶园农药减施增效、降低茶叶中农药残留，提高茶叶质量安全水平。

本项目通过进行茚虫威水解、光解试验研究，借助UPLC-QTOF-MS技术筛查、文献报道并委托合成相关降解产物进行比对验证，明确了水解、光解降解产物种类（IN-JT333、IN-MF014、IN-MK638、IN-KT413、IN-MU716、IN-P0036）和规律；考察了不同色谱柱对茚虫威及其手性结构代谢物的对映体分离效果，利用UPLC-QTOF-MS和UPLC-MS/MS建立了茶鲜叶、红茶、绿茶和茶汤中茚虫威及其7种降解产物对映体、外消旋体残留分析方法，方法能够满足残留分析方法要求和后续样本检测的需要；利用手性制备色谱柱拆分茚虫威对映体获得对映单体，对茚虫威对映单体进行田间喷施残留试验，采集茶鲜叶进行红茶、绿茶加工，冲泡茶汤，借助UPLC-MS/MS技术定性定量分析研究了茚虫威对映体在茶叶生长-加工-冲泡中的代谢产物有IN-JT333、IN-MK638、IN-MF014和IN-KG433及其迁移转化差异规律，进一步验证R-indoxacarb存在转化为S-indoxacarb趋势，首次在植物源样本中发现代谢物IN-JT333但在茶汤中未检出，明确了绿茶的杀青、揉捻和红茶的揉捻、发酵是影响茚虫威手性对映体及降解产物含量变化最显著的工艺，在冲泡过程中，绿茶中R-indoxacarb和S-indoxacarb的浸出率均略高于红茶但均不超过10%，而代谢物IN-KG433、IN-MK638和IN-MF014的最大浸出率分别达88.2%、89.9%和94.1%；首次研究分析了茚虫威对映体对茶尺蠖的杀虫活性差异和对蜜蜂的毒性差异，明确对映体之间对茶尺蠖的活性差异和对蜜蜂的毒性差异均为S-indoxacarb > indoxacarb (3S + 1R) >> R-indoxacarb；利用计算机毒性预测软件进行茚虫威代谢物的毒性预测，明确降解产物的毒性IN-KG433 > indoxacarb > IN-JT333 > IN-MK638 > IN-MF014，其中IN-KG433的毒性和浸出率均高于茚虫威，在冲泡过程中很容易从茶叶转移到茶汤中，所幸的是在茶叶中的残留量不高，不会导致饮用安全风险。项目总结果为指导茶园手性农药合理施用和有效开展茶园手性农药风险评估提供了参考借鉴。