Design, Synthesis, and Study of the Dual Action Mode of Novel N-Thienyl-1,5-disubstituted-4-pyrazole Carboxamides against Nigrospora oryzae

Due to the single target but extensive application of commercialized succinate dehydrogenase inhibitors (SDHIs), resistance problems have gradually become apparent in recent years. To solve this problem, a series of novel N-thienyl-1,5-disubstituted-1H-4-pyrazole carboxamide derivatives were designed and synthesized in this work based on the active skeleton 5-trifluoromethyl-4-pyrazole carboxamide. The bioassay results indicated that some target compounds exhibited excellent in vitro Antifungal activities against the eight phytopathogenic fungi tested. Among them, the EC₅₀ values of T₄, T₆, and T₉ against Nigrospora oryzae 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 studies revealed that T₆ not only significantly inhibited the growth of N. oryzae mycelia but also effectively hindered spore germination and germ tube elongation. Morphological studies using scanning electron microscopy (SEM), fluorescence microscopy (FM), and transmission electron microscopy (TEM) found that T₆ could 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 T₆ against succinate dehydrogenase (SDH) was 7.2 mg/L, lower than that of the commercialized SDHI penthiopyrad (3.4 mg/L). 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. Thus, this study provides a new idea for a strategy to delay resistance and diversify the structures of SDHIs.

5-disubstituted-1H-4-pyrazole carboxamides; N-thienyl-1; antifungal activity; cell membrane integrity; dual mode of action against N. oryzae; succinate dehydrogenase inhibitors.