手性农药对映异构体对葡萄酒发酵及品质影响的差异研究

The grapes were usually suffered from several diseases during growth, especially fungal infection such as downy mildew, white rot, gray mold and so on. In this case, pesticides are inevitably applied to control these diseases until the grapes ripen. Worse more, it has been demonstrated that the pesticide residues may remain in the products obtained from these polluted grapes, such as wine, which might cause potential risk in food safety and human health. On the other hand, it was found that some of the pesticide residues had certain influence on the wine fermentation process.. However, among the common pesticides, about 25% of them are chiral, which contains more than one enantiomer, and they are usually applied and released into environment as racemic form. Interestingly, it has been demonstrated by many researchers that the enantiomers of chiral pesticides usually have differences in activity, toxicity, degradation and environmental behaviors. In the previous study, it was reported that stereoselective dissipation of epoxiconazole occured in the grape, and it was also observed that diclofop-methyl showed enantioselective degradation during alcohol fermentation process. Consequently, it is of great significance to investigate the influence of the pesticide residues to wine yeast at enantiomer level.. This project aims to reveal the metabolic stress of the fungicide enantiomers to the wine yeast. Three typical triazole fungicides (epoxiconazole, hexaconazole, vinclozolin and metalaxyl) are chosen for study in this experiment.The technology of HPLC-MS-MS-CSP will be applied in the analysis of the enantiomers and the metabolite of the chiral pesticides.The effect of these enantiomers on the enzymes in the yeast will also be studied. An analytical method based on GC/HPLC-MS-MS will be established for the metabonomics study of common fermentation metabolites. Two strains of yeast(Saccharomyces cerevisiae and Kloeckera apiculata) are applied to study the metabolic stress of the selected pesticides. With these results, the mechanism of the metabolic stress can be clear. . With the study of this project, the metabolic stress of the fungicide enantiomers will be clarified, and the difference of each enantiomer can also be clear. These results will also provide more detailed technical data for the evaluation of the potential risk of red wine quality.

大量研究发现农药残留对葡萄酒酵母具有抑制作用，并影响葡萄酒发酵过程，但在对映异构体水平上，从分子生物学角度考察手性农药对酵母影响的研究甚少。本项目以杀菌剂氟环唑、己唑醇、乙烯菌核利和甲霜灵为研究对象，考察其在两种不同葡萄酒酵母发酵葡萄酒过程中的立体选择性降解情况及代谢物生成情况，并考察不同对映异构体对发酵过程中酵母生长、酵母酶活力及发酵产物的影响，尤其是不良代谢产物的变化。在此基础上，结合酶学试验结果，探讨手性农药对葡萄酒品质影响的作用机制。该项研究的开展，可全面、深入地评价手性农药对映异构体对葡萄酒酿造的影响，为葡萄酒品质及安全风险评估提供相关理论依据和更多的数据支持。另外，相关结果可为高品质葡萄酒生产原料中的农药残留的控制提供有效指导数据，为葡萄酒产业的长足发展奠定基础。

农药残留不仅影响食品的安全性，同时可能会影响食品加工过程，例如生物发酵过程。本项目主要开展了以下研究工作：（1）手性农药己唑醇、氟环唑、甲霜灵在葡萄酒酿造过程中的立体选择性降解研究，己唑醇、氟环唑及甲霜灵在葡萄酒酿造过程中的立体选择性降解及残留情况；（2）项目采用代谢组学手段，利用现代仪器分析技术，建立了葡萄酒中常见代谢物（氨基酸、醇类化合物、酯类化合物等）的分析方法，利用代谢组学和统计学分析方法，完成了己唑醇、粉唑醇、苯醚甲环唑、戊唑醇、丙环唑残留对葡萄酒品质的影响研究。（3）由于葡萄酒酿造过程中，微生物菌群相对简单，供试农药降解趋势不明显，且酿造周期较长，因此，为充分考察酒类发酵过程中，手性农药残留变化情况，项目组同时开展了腈菌唑、己唑醇、氟环唑、多效唑在黄酒酿造过程中的立体选择性降解及其归趋研究，并考察了腈菌唑和己唑醇对映体对酿酒酵母及米曲霉中相关酶的影响情况。. 研究结果表明，杀菌剂残留会对葡萄酒酵母产生一定的影响，包括对酵母的生长、相关酶活力以及代谢产物。因此，不能以食用葡萄中的农药残留限量来控制葡萄酒原料，有必要对葡萄酒酿造用葡萄中的农药残留限量进行进一步的严格限定，以保证葡萄酒的安全性及其品质。另外，发酵过程中，微生物种群情况会对农药降解影响较大，种群丰富，可有利于农药的降解。. 该项研究的开展，可全面、深入地评价手性农药对映异构体对葡萄酒酿造的影响，为葡萄酒品质及安全风险评估提供相关理论依据和更多的数据支持。另外，相关结果可为高品质葡萄酒生产原料中的农药残留的控制提供有效指导数据，为葡萄酒产业的长足发展奠定基础。