This study aimed to develop novel and highly effective antifungal agents to control key plant diseases. Based on the 2-chloronicotinohydrazide core structure, 22 new nicotinohydrazide derivatives were designed and synthesized by optimizing phenoxy substitution sites. Antifungal evaluation revealed that this series of compounds possess highly effective broad-spectrum fungicidal activity. Among them, compound J15 exhibited particularly outstanding inhibitory activity against Rhizoctonia solani (EC50 = 0.13 μg/mL), and its control efficacy in both detached leaf assays and rice pot tests was superior to that of boscalid. Molecular docking revealed that J15 enhances its binding affinity to succinate dehydrogenase (SDH) through a hydrophobic interaction network formed by key amino acid residues (ILE40C, ARG43C). This result aligns with J15's IC50 value being 1.8-fold lower than that of boscalid. Mechanistic studies demonstrated that J15 exerts its fungicidal action by inhibiting SDH, thereby inducing reactive oxygen species (ROS) accumulation, and cell membrane damage, as evidenced by mitochondrial membrane potential depolarization and increased PI uptake. Regarding safety, J15 has minimal effects on the viability of human lung cells (BEAS-2B) and human liver cells (THLE-2) within the effective antifungal concentration range (0.28-3.4 μM) and agricultural-related concentrations (<1 μM), with cell viability remaining above 98 %. These findings position J15 as a promising lead for next-generation, dual-mechanism SDHIs with improved resistance profiles and safety.
