Adsorption and Structural Change of β-Lactoglobulin at the Diacylglycerol−Water Interface

Diacylglycerol (DAG)/water and triacylglycerol (TAG)/water emulsions were prepared using β-lactoglobulin (β-LG) as an emulsifier. The oil phase (20% in emulsion) was mixed with β-LG solution (1% β-LG in water, pH 7) to prepare the emulsions. A fine oil-in-water emulsion was produced from both DAG and TAG oils. The interfacial protein concentration of the TAG emulsion was higher than that of the DAG emulsion. The ζ potential of the DAG oil droplet was higher than that of the TAG oil droplet. The front-surface fluorescence spectroscopy results revealed that tryptophan residues in β-LG moved to the more hydrophobic environment during the adsorption of protein on the oil droplet surfaces. Changes in secondary structure of β-LG during the adsorption were determined by FT-IR spectroscopy. Decreases in the β-sheet content concomitant with increases in the α-helix content were observed during the adsorption to the oil droplets, and the degree of structural change was greater for β-LG in the TAG emulsion than in the DAG emulsion, indicating the increased unfolding of adsorbed β-LG on the TAG oil droplet surface. Results of interfacial tension measurement supported this speculation, that is, the increased unfolding of the protein at the TAG−water interface. Trypsin- and proteinase K-catalyzed proteolysis was used to probe the topography of the adsorbed β-LG on the oil droplet surface. SDS−PAGE analyses of liberated peptides after the proteolysis indicated the higher susceptibility of β-LG adsorbed on the DAG oil droplet surface than on the TAG oil droplet surface. On the basis of all the results, we discussed the conformation of the adsorbed β-LG on the two oil droplet surfaces.