BACKGROUNDThe potential application of vanillin as a fungicide has garnered significant attention in the agricultural product market and food industries. Consequently, a novel series of vanillin derivatives containing thiazole and hydrazone fragments were strategically designed, synthesized, and evaluated for their antifungal activity against six representative plant phytopathogenic fungi.RESULTSIn the in vitro antifungal assay, some title vanillin derivatives showed good antifungal activity against Botrytis cinerea, Fusarium solani, and Magnaporthe grisea. Significantly, compound 4a exhibited remarkable broad‐spectrum fungistatic potency and displayed the most potent antifungal activity against B. cinerea and F. solani, with half maximal effective concentration (EC50) values of 1.07 and 0.78 μg/mL, respectively, substantially surpassing the efficacy of the commercial fungicide hymexazol and comparable to carbendazim. In addition, compound 4k selectively inhibited M. grisea with the lowest EC50 value of 7.77 μg/mL. The in vivo antifungal assay revealed that compound 4a exhibited superior protective efficacy against B. cinerea compared to carbendazim. At the same time, scanning electron microscopy (SEM), transmission electron microscopy (TEM), and fluorescent dye staining showed compound 4a exerted its fungicidal activity by damaging the cell wall, cell membrane and mitochondria of B. cinerea.CONCLUSIONModification of vanillin through electron‐withdrawing group substitution and hydrophobic substitution, followed by condensation with thiazole‐4‐carbohydrazide, could result in highly active antifungal derivatives. Among them, compound 4a, which exhibited excellent inhibitory activities in vitro and in vivo, could potentially serve as a lead compound. © 2024 Society of Chemical Industry.