Enhancement of the interfacial behaviors of β-lactoglobulin fibrils of different morphology by calcium ions

Protein self-assembled fibrils have attracted extensive attention due to their unique properties, but they exhibit high sensitivity to salt ions. This study investigated the effect of Ca²⁺ on the interfacial rheological behaviors of long β-lactoglobulin fibrils (LFblgs, long and straight fibrillar structures with contour lengths of 700–2500 nm and diameters of 9–16 nm) and short β-lactoglobulin fibrils (SFblgs, curled worm-like fibrils with lengths of 100–320 nm and diameters of 2.5–7 nm), which were prepared by modulating solvent polarity. The results showed that the addition of Ca²⁺ decreased the ζ-potential values of both LFblgs and SFblgs, which was attributed to salt bridge formation between Ca²⁺ and carboxyl groups on the protein fibrils. Rheological measurements further confirmed this result, showing that Ca²⁺ enhanced the viscoelastic moduli of both fibril systems at appropriate concentrations and promoted the formation of a dense but weak semi-flexible network, as indicated by a power-law relationship between Ca²⁺ concentration and the elastic shear modulus G′ of the gel. Although Ca²⁺ hindered fibril adsorption to the O/W interface, it increased the interfacial pressure and enhanced the viscoelasticity of the protein fibril layer by forming an inhomogeneous network at the O/W interface, as demonstrated by interfacial dilatational, shear and particle tracking rheology. The study reveals the mechanism of Ca²⁺ enhanced interfacial behavior of β-lactoglobulin fibrils through the use of various analytical techniques, providing valuable insights for the application of β-lactoglobulin fibrils in the food industry.