BACKGROUNDBiofilm plays a crucial role in Pseudomonas syringae pv. tomato (Pst) infection. We identified (1R,4S)‐(−)‐fenchone (FCH) as the most potent antibiofilm agent against Pst among 39 essential oil compounds. Subsequently, we synthesized a series of FCH oxime ester and acylhydrazine derivatives to explore more potent derivatives.RESULTSII3 was screened out as the most potent derivative, exhibiting a minimal biofilm inhibitory concentration of 60 μg mL−1 and a lowest concentration with maximal biofilm inhibition (LCMBI) of 200 μg mL−1, lower than those of FCH (80 and 500 μg mL−1, respectively). II3 and FCH showed minimum inhibitory concentration values >1000 μg mL−1 and similar maximal biofilm inhibition extents of 48.7% and 49.5% at their respective LCMBIs, respectively. Meanwhile, neither of them influenced cell viability or the activity of metabolic enzymes at their respective LCMBIs. II3 at its LCMBI significantly reduced biofilm thickness, extracellular polysaccharide content, and pectinase and cellulase production indices. In vivo assay results indicated that II3 could preventatively reduce the bacterial contents in tomato leaves at its LCMBI, and when combined with kasugamycin (KSG) (10 μg mL−1), II3 achieved the same level of bacterial reduction as the sole application of KSG (70 μg mL−1), thereby reducing the required dosage of KSG. Mechanistic studies demonstrated that II3 can down‐regulate biofilm‐related genes and inhibit PsyR/PsyI quorum sensing system, which differs from the bactericidal mechanisms.CONCLUSIONThese results underscore the potential of II3 as an antibiofilm agent for the control of Pst or FCH as a promising natural candidate for future in‐depth optimization. © 2024 Society of Chemical Industry.