Raphani Semen Extract Targets the las Quorum Sensing System to Suppress Pseudomonas aeruginosa Biofilm and Signal Molecule Production

ABSTRACT

Raphanus sativus L. seeds (Raphani Semen, RS) are traditionally used in Chinese medicine for respiratory and gastrointestinal disorders. Pseudomonas aeruginosa ( P . aeruginosa ) biofilm formation and quorum sensing (QS)‐regulated behaviors contribute significantly to its persistence and resistance, highlighting the need for novel QS‐targeting agents. This study aimed to investigate whether RS extract (RSE) and its characteristic constituents sulforaphane (SFA) and sulforaphene (SFE) inhibit P. aeruginosa biofilm and virulence by targeting specific QS pathways. Using sub‐inhibitory concentrations of RSE (≤ 256 μg/mL), SFA and SFE (12.5–100 μM), we evaluated their effects on biofilm formation, bacterial motility, virulence factor production, and signal molecule levels, combined with fluorescent reporter assays, qRT‐PCR, and UPLC‐Q‐Orbitrap‐MS/MS for mechanistic and phytochemical analysis. Results showed that RSE suppressed biofilm formation by 50.93%, impaired motility, and reduced 3‐oxo‐C12‐HSL production. It downregulated key QS genes ( lasR , lasA , lasB , pqsA , and pqsR ) and inhibited lasB / pqsA reporter activity. Phytochemical analysis identified 71 components, and subsequent validation showed that SFA and SFE replicated these effects and exhibited broader QS inhibition across the las , pqs , and rhl systems. SFA (100 μM) demonstrated particularly strong efficacy, suppressing 3‐oxo‐C12‐HSL by 74.29%. Molecular docking indicated that SFA and SFE can bind to LasB, RhlA, and PqsA, albeit with modest affinities. These findings demonstrate that RSE and its active components, SFA and SFE, are potent QS inhibitors that attenuate P. aeruginosa pathogenicity primarily by disrupting the las system, suggesting their potential as natural anti‐biofilm agents that may target multiple components of the QS system.