Interaction Dynamics of Liposomal Fatty Acids with Gram-Positive Bacterial Membranes

The increasing prevalence of antibiotic-resistant bacteria has driven the need for alternative therapeutic strategies, with liposomal fatty acids (LipoFAs) emerging as promising candidates due to their potent antibacterial properties. Despite growing interest, the detailed biophysical interactions between LipoFAs and bacterial membranes remain underexplored. In this study, we systematically investigate the mechanistic interactions of liposomal linolenic acid (LipoLNA), linoleic acid (LipoLLA), and oleic acid (LipoOA) with model Gram-positive bacterial membranes using quartz crystal microbalance with dissipation (QCM-D) and fluorescence microscopy. QCM-D analysis revealed that LipoOA displayed the highest rate of membrane fusion, followed by LipoLLA and LipoLNA. Fluorescence microscopy highlighted distinct morphological changes induced by each LipoFA: LipoLNA generated large membrane buds, LipoLLA formed smaller dense protrusions, and LipoOA caused rapid incorporation with uniform dense spots. Furthermore, fluorescence recovery after photobleaching (FRAP) demonstrated that LipoLNA significantly enhanced lipid mobility and membrane fluidity, as confirmed by Laurdan generalized polarization measurements. The extent of unsaturation in LipoFAs was found to play a critical role in their interaction mechanism, with higher degrees of unsaturation inducing greater local curvature stress, increased membrane permeability, and substantial ATP leakage, ultimately leading to improved bactericidal activity. Notably, liposomal formulations exhibited enhanced biocompatibility compared to free fatty acids. These findings provide valuable mechanistic insights into how LipoFAs perturb bacterial membranes, supporting their potential application as alternative antibacterial agents.