Isolation and purification of serum and interfacial peptides of a trypsinolyzed β-lactoglobulin oil-in-water emulsion

Information on the conformation of proteins adsorbed to an oil–water interface is usually determined by following the time course of enzymatic hydrolysis of the protein in an oil-in-water emulsion. Unlike previous works reported in the literature, the research presented in this paper provides information on which peptides are actually in contact with the lipid bilayer (interfacial peptides) and those segments that project into the aqueous phase (serum peptides). In order to achieve this classification of peptides, we present a method to separate serum peptides from interfacial peptides by initial centrifugation steps followed by reversed-phase high-performance liquid chromatography. The effectiveness of the method was ascertained by performing proteolysis on β-lactoglobulin adsorbed to an oil-water interface in a soybean oil-water emulsion. It was found that more peptides are qualitatively and quantitatively found adsorbed to the oil–water interface as compared to peptides released into the serum.