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標題: 牛筋草對伏寄普之抗性生理機制研究-代謝解毒反應
Resistance mechanism of goosegrass (Eleusine indica (L.) Gaertn.) resistant to fluazifop-butyl - metabolic detoxification
作者: Wan-Ting Lin
關鍵字: herbicide;resistance;weed;metabolism;fluazifop-P-butyl;除草劑;抗性;雜草;代謝;伏寄普
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禾草類除草劑(graminicides)屬於選擇性早期萌後型除草劑,此?除草劑之作用機制為抑制催化脂肪酸合成第一步驟之酵素-乙醯輔?A羧化酵素(acetyl-CoA carboxylase; ACCase; EC之活性,進而抑制脂肪酸合成,破壞細胞膜之完整性。本研究為了解牛筋草[Eleusine indica (L.) Gaertn.]對ACCase抑制型除草劑伏寄普(fluazifop-P-butyl)產生抗性之機制,分析牛筋草抗、感性生物型(biotype)植株對於該藥劑之吸收、轉運以及代謝之差異。結果顯示在藥劑吸收方面,牛筋草抗感性生物型之間並無顯著差異,而在轉運方面,抗性生物型雖然對於伏寄普具有較高之轉運能力,能將20% 14C-fluazifop-P-butyl及其代謝物轉運至上位葉,但於伏寄普處理後7天,處理葉與上位葉存在之fluazifop-buty約90%已轉變成伏寄普酸(fluazifop acid)及其代謝物(metabolites),尤其抗性生物型具有較高比例之代謝物,因此推測代謝能力差異可能是造成抗性之原因之一。
根據質譜儀分析結果顯示,於正離子模式下出現m/z 512, 432, 423, 415, 314以及160等6個訊號以及在負離子模式下出現m/z 788, 623, 593以及162等4個訊號為抗、感生物型共有之代謝物,但除m/z 162之外,在抗性生物型中均表現較強訊號。正離子模式下於出現m/z 202.180、219.012、255.944、288.963、310.156與340.865等6個訊號為抗性生物型中獨有之代謝物, m/z 333.011、525.977、631.027、647.003、688.030、827.067、941.060、989.120、182.034、191.039、200.044、214.918、265.023、322.937、328.887、338.343、352.948、365.107、374.038、381.080與394.920等21個訊號為感性生物型中獨有之代謝物,顯示抗、感生物型有不同的代謝物剖面(profile)。從發現之代謝物出現2-[4-(5-trifluoromethyl-2-pyridyloxy)phenoxy] propanol、5-trifluoromethyl-2-hydroxy-pyridine與5-trifluoromethyl-2- pyridone,推測可能與細胞色素P450還原?(cytochrome P450 reductase, CYP; EC參與之還原反應(reduction)有關。因此,進一步分析細胞色素P450還原?活性。
酵素活性分析結果顯示,經伏寄普處理後抗性生物型具有較高之細胞色素P450還原?活性,推測在牛筋草抗性生物型中伏寄普(fluazifop-P-butyp)經水解作用(hydrolysis)形成伏寄普酸(fluazifop acid)後,可能透過細胞色素P450還原?行還原反應(reduction)形成2-[4-(5-trifluoromethyl-2-pyridyloxy)phenoxy]propanol,可能再轉變成4-(5-trifluoromethyl-2-puridyl)oxyphenol之後,最後再透過細胞色素P450還原?行還原反應(reduction)形成5-trifluoromethyl-2-hydroxy- pyridine與5-trifluoromethyl-2-pyridone。

Fluazifop-P-butyl, a selective graminicide, is widely used to control annual grass weeds. The action mechanism of this herbicide is to inhibit the activity of acetyl-CoA carboxylase (ACCase; EC, which is responsible for the lipid biosynthesis. In order to explore the resistance mechanism, resistant (R)- and susceptible (S)-biotype of goosegrass [Eleusine indica (L.) Gaertn.] treated with 14C-fluazifop-P-butyl were studied. Although no significant difference of herbicide uptake between R- and S-biotype, and even the higher translocation rate of 14C-fluazifop and its metabolites in R-biotype was found, it is suggested that the herbicide metabolism is involved in resistance mechanism due to the more polar metabolites were found in R-biotype.
According to the results of mass spectrum analysis, six signals including m/z 512, 432, 423, 415, 314 and 160 under POS scanning mode, and four signals including m/z 788, 623, 593 and 162 under NEG scanning mode appeared in both R- and S-biotype. These signals had stronger intensities in R-biotype, except m/z 162. Interestingly, six signals including m/z 202.180, 219.012, 255.944, 288.963, 310.156 and 340.865, only appeared in R-biotype, and twenty-one signals including m/z 333.011, 525.977, 631.027, 647.003, 688.030, 827.067, 941.060, 989.120, 182.034, 191.039, 200.044, 214.918, 265.023, 322.937, 328.887, 338.343, 352.948, 365.107, 374.038, 381.080 and 394.920 only appeared in S-biotype, suggesting different metabolite profile between R- and S-biotype. In this metabolite study, the generation of 2-[4-(5-trifluoro- methyl-2-pyridyloxy)phenoxy] propanol, 5-trifluoromethyl-2-hydroxy- pyridine and 5-trifluoromethyl-2-pyridine suggested that certain reduction reaction might be happened. Therefore, we attempted to clarify the role of cytochrome P450 reductase (CYP; EC in fluazifop metabolism.
Coupled with the study on the activity of cytochrome P450 reductase, it is suggests that fluazifop-P-butyl in goosegrass was hydrolyzed to fluazifop acid, and then reduced to 2-[4-(5-trifluoromethyl-2-pyridyloxy) phenoxy] propanol by cytochrome P450 reductase. Subsequently, 4-(5-trifluoromethyl-2-puridyl)oxyphenol was formed, and both 5-tri- fluoromethyl-2-hydroxy-pyridine and 5-trifluoromethyl-2-pyridone were formed through reduction catalyzed by cytochrome P450 reductase.
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