Study of triglyceride‐ protein interaction using a microemulsion‐ filtration method

Interactions between simple triglycerides and a variety of native and chemically modified proteins were studied. A new procedure was developed to evaluate hydrophobic binding of proteins at oil/water interfaces. The method involves emulsifying a triglyceride ultra‐sonically in water and equilibrating the microemulsion with a protein solution. The protein adsorbs to the surfaces of the globules as a stabilizing interfacial layer. Relative stability of different emulsions is evaluated under standard conditions of partition with a poly‐carbonate membrane filter. Either gas liquid chromatography or a radioactive technique is used to determine the amount of trigly‐ceride stabilized by protein through hydrophobic binding. The procedure was used to compare interactions of six different trigly‐cerides with bovine serum albumin (BSA) and interactions between trioctanoin and 26 different native and chemically modified pro‐teins. In general, an increase in molecular weight of saturated trigly‐cerides or an increase in double bonds of unsaturated triglycerides reduced hydrophobic binding. The order of binding of trioctanoin for the series of proteins (on a weight basis) compared to BSA taken as 100%, ranged from 160% for casein to 1% for lysozyme. Addition of nonpolar residues, including methyl, cyclohexyl, cyclopentyl and benzyl groups, to casein increased binding by 20–30%. Effects of protein mixtures, and changes in pH and ionic strength also were studied. The results demonstrate that interactions between a wide variety of triglycerides and proteins can be investigated by the microemulsion‐filtration method.