pH-Dependent charge regulation and ion-specific supramolecular organization in surfactin-iturin-rich bioamphiphile...

Surfactin- and iturin-containing biosurfactants are commonly treated as anionic lipopeptides, although their headgroups contain titratable acidic and, where available, basic functional groups that can influence interfacial charging and supramolecular organization. Here, we investigate surfactin-iturin-rich bioamphiphile (BAP) preparations enriched in surfactin and iturin and show that they display pronounced pH-dependent charge regulation in bulk solution and at interfaces. Potentiometric titration revealed two apparent pK_a values, while surface tension measurements identified a critical micelle concentration of approximately 1 mg mL^-1. Small-angle X-ray scattering indicated the formation of supramolecular aggregates, including elongated assemblies on the submicrometre scale coexisting with smaller scattering domains. To probe local interfacial acid-base properties, we used 1-hydroxy-2-naphtoic acid as a spectroscopic reporter and found that the apparent interfacial pH differs from the bulk environment and responds sensitively to ion identity. Combined zeta-potential, dynamic light scattering, and apparent interfacial pH measurements revealed strong ion-specific effects under acidic conditions, including changes in apparent aggregate size and surface charging that do not follow a simple Hofmeister trend. Instead, the results support a picture in which charge regulation, ion hydration, local aggregate polarity, and ion polarizability collectively govern the interfacial organization of these biosurfactant assemblies. These findings reframe surfactin-iturin-rich mixtures as responsive bioamphiphile systems and provide a physicochemical basis for understanding and tuning their behavior in complex aqueous environments.