Structural rearrangement of beta-lactoglobulin at different oil-water interfaces and its effect on emulsion stability

Understanding the factors that control protein structure and stability at the oil-water interface continues to be a major focus to optimize the formulation of protein stabilized emulsions. In this study, a combination of synchrotron radiation circular dichroism spectroscopy (SRCD), front-face fluorescence spectroscopy and dual polarization interferometry (DPI) were used to characterize the conformation and geometric structure of beta-lactoglobulin (beta-Lg) upon adsorption to two oil-water interfaces: a hexadecane-water interface and a tricaprin-water interface. The results show that upon adsorption to both oil-water interfaces, beta-Lg went through a beta-sheet to alpha-helix transition with a corresponding loss of its globular tertiary structure. The degree of conformational change was also a function of oil phase polarity. The hexadecane oil induced a much higher degree of non-native alpha-helix compared to the tricaprin oil. In contrast to beta-Lg conformation in solution, the non-native alpha-helical-rich conformation of beta-Lg at the interface was resistant to further conformational change upon heating. DPI measurements suggest that beta-Lg formed a thin dense layer at emulsion droplet surfaces. The effect of high temperature and the presence of salt on these beta-Lg emulsions were then investigated by monitoring changes in zeta-potential and particle size. In the absence of salt, high electrostatic repulsion meant beta-Lg stabilized emulsions were resistant to heating to 90 oC. Adding salt (120 mM NaCl) before or after heating led to emulsion flocculation due to the screening of the electrostatic repulsion between colloidal particles. This study has provided insight into the structural properties of proteins adsorbed at the oil-water interface and has implications in the formulation and production of emulsions stabilized by globular proteins.