Natural deep eutectic solvents as environmentally compatible agents for biofilm inhibition and disruption

Natural deep eutectic solvents (NADES) have recently emerged as promising 'green' antibiofilm agents due to their ability to solubilize biological macromolecules. In this study, three chemically distinct NADES formulations were evaluated against Pseudomonas fluorescens WCS365 and Staphylococcus epidermidis ATCC 35984. The tested formulations choline chloride-lactic acid (CCLA), choline chloride-urea (CCU), and choline chloride-xylitol (CCX) were assessed for their effects on planktonic growth, biofilm formation, and mature biofilms. All NADES showed moderate inhibition of planktonic growth, while biofilm formation was significantly reduced in a formulation- and species-dependent manner, with CCLA displaying the strongest activity. Treatment of pre-formed 24 h biofilms resulted in partial but significant biomass reduction, reaching up to ∼69% for P. fluorescens and ∼51% for S. epidermidis. Confocal microscopy confirmed pronounced structural disruption of mature biofilms following NADES exposure, particularly for CCLA. These findings highlight the potential of organic acid-based NADES as biocides, which are more environmentally compatible than conventional biocides.