Design,Synthesis And Biological Activity Of N-Thiophene-1,5-Disubstituted-4-Pyrazole Amide Derivatives

Plant pathogenic fungi pose a huge threat to global food security,and fungicides are our most powerful weapon against plant pathogenic fungi.Succinate dehydrogenase inhibitors（SDHIs）are a class of fungicides that inhibit the respiration of pathogenic fungi by inhibiting the activity of succinate dehydrogenase in the mitochondrial respiratory electron transport chain and destroying the tricarboxylic acid cycle.SDHIs play an important role in the fungicide market due to their high efficiency,internal absorption and broad spectrum.However,due to the single and intensive use of SDHIs targets,resistance problems have gradually emerged in recent years,resulting in SDHIs being classified as having a medium-high resistance risk by the Fungicide Resistance Action Committee（FRAC）.Therefore,the development of fungicides with multiple modes of action can not only delay the development of drug resistance,but also provide ideas for the development of new fungicides.In this paper,based on the active skeleton of“1-substituted-5-trifluoromethyl-4-pyrazole amide”discovered by our group in the early stage,30“N-thiophene-1,5-disubstituted-4-pyrazole amide derivatives”were designed and synthesized by introducing different substituents at the“1”and“5”positions of the pyrazole ring and introducing thiophene groups at the amino site.The biological activity tests showed that the EC₅₀ values of compounds T₄,T₆ and T₉ were 5.8,1.9 and 5.5 mg/L,respectively.The in vivo protective and curative activities of 40 mg/L T₆ against rice infected with N.oryzae were 81.5%and 43.0%,respectively.Further mechanism studies showed that compound T₆ not only significantly inhibited the growth of N.oryzae mycelia but also effectively hindered spore germination and germ tube elongation.Morphological studies using fluorescence microscopy（FM）,scanning electron microscopy（SEM）and transmission electron microscopy（TEM）found that T₆could affect the mycelium membrane integrity by increasing cell membrane permeability and causing peroxidation of cellular lipids,and these results were further verified by measuring the malondialdehyde（MDA）content.The IC₅₀ value of compound T₆ against succinate dehydrogenase（SDH）was 7.2 mg/L,indicating that the compound had a significant inhibitory effect on SDH.Further,ATP content detection and the results after docking T₆ and penthiopyrad suggested that T₆ was a potential SDHI.These studies demonstrated that active compound T₆ could both inhibit the activity of SDH and affect the integrity of the cell membrane at the same time via a dual action mode,which is different from the mode of action of penthiopyrad.Hereby,this paper provides a new idea and direction for the development of SDHIs.