Discovery of Novel Isophorone Derivatives as Potential Succinate Dehydrogenase Inhibitors: Design, Synthesis, Antifungal Evaluation, and Action Mechanism

In this work, a total of 45 novel isophorone derivatives were designed, synthesized, and evaluated for antifungal activity against six phytopathogenic fungi. Some target compounds displayed remarkable and broad-spectrum antifungal activities <i>in vitro</i> against tested phytopathogenic fungi. Among them, compound <b>A2</b> exhibited excellent antifungal activity against <i>Rhizoctonia solani</i>, <i>Sclerotinia sclerotiorum</i>, <i>Valsa mali</i>, <i>Botrytis cinerea</i>, <i>Gibberella zeae</i>, and <i>Physalospora piricola</i>, with the corresponding EC₅₀ values of 0.133, 0.258, 0.428, 0.519, 1.29, and 1.51 μg/mL, respectively. Additionally, results from <i>in vivo</i> experiments indicated that compound <b>A2</b> could function as a novel antifungal candidate for safeguarding agricultural crops against fungal infections. During the investigation into the antifungal mechanism, the cell membrane permeability and propidium iodide (PI) staining experiments demonstrated that compound <b>A2</b> could destroy the cell membrane structure and increase the permeability of the cell membrane. Findings from microscopic observations, in tandem with mitochondrial membrane potential (MMP) detection, revealed that compound <b>A2</b> severely damaged the structural integrity of cells. Concurrently, compound <b>A2</b> decreased the MMP, thereby inducing cell apoptosis and inhibiting the normal growth of mycelia. Moreover, results from succinate dehydrogenase (SDH) enzyme assays, molecular dynamics (MD) simulations, and molecular docking experiments further indicated that compound <b>A2</b>, Thifluzamide, and boscalid might have similar mechanisms of action and binding modes with SDH. Finally, to investigate the structural basis for differences in bioactivity, the frontier molecular orbitals and molecular electrostatic potential were calculated. The outcomes of this work significantly contribute to further research aimed at agricultural plant disease control.