Preharvest Plai Oil Application in Hydroponic Kale for Producing Fresh Salad Leaves with Antimicrobial Activity Against <i>Listeria monocytogenes</i> : Possible Mechanism of Action

A novel preharvest method for producing antimicrobial kale in hydroponic systems was developed using a low-concentration plai oil nanoemulsion. The nanoemulsion was applied to the hydroponic solution at concentrations of 5, 10, 15, and 20 μL/mL for 12 h prior to harvest. Antimicrobial activity against <i>Listeria monocytogenes</i> was evaluated in both treated and control kale samples. In addition, bioactive compounds, volatile oil residues, and the potential mechanism of antimicrobial action were investigated. Application of 10 μL/mL plai oil resulted in a reduction of <i>L. monocytogenes</i> by more than 5 log₁₀ colony-forming units per gram and completely inhibited disease incidence during storage at 4°C for at least 7 days. Although higher concentrations (15 and 20 μL/mL) also reduced bacterial counts, increased wilting was observed in the leaves. In contrast, the control group developed visible disease symptoms within 3 days of storage. Based on microbial suppression and visual quality, 10 μL/mL was identified as the optimal concentration. The preharvest treatment significantly enhanced the levels of bioactive compounds and antioxidant activity compared with control kale. Volatile constituents of plai oil, including thujene (35%), terpinene (21%), pinene (15%), and terpinene-4-ol (14%), were detected in treated kale. The antimicrobial mechanism was associated with membrane disruption, as evidenced by porous and shriveled cell walls observed in <i>L. monocytogenes</i>, along with decreased pH, increased conductivity, and elevated protein and amino acid leakage after 18 h of inoculation. Biofilm formation by <i>L. monocytogenes</i> on kale was also significantly inhibited. This preharvest strategy offers a practical and effective method for enhancing microbial safety and extending the shelf life of hydroponically grown kale.