INTERACTION OF SUCROSE ESTERS WITH SKIM MILK-PROTEINS AS CHARACTERIZED BY AFFINITY-CHROMATOGRAPHY

The interaction of sucrose ester, P-1670 (70% pure palmitic acid monoester), with milk proteins was examined by affinity chromatography. The ester was covalently bound to 3-aminopropyl controlled-pore glass by periodate oxidation of the carbohydrate to aldehyde, formation of the Schiff base, followed by reduction to secondary amine. In phosphate buffer, pH 7, alpha-LA did not appear to interact with sucrose ester, but beta-LG exhibited binding. However, casein micelles were very tightly bound, and caseins could only be eluted following addition of urea. On application of skim milk, the strengths of the interactions, as reflected by their elution volumes, decreased in the order: alpha(s)-CN, kappa-CN, beta-CN, beta-LG, and alpha-LA. Micelles dissociate after addition of urea, allowing individual caseins to interact with sucrose ester moieties in the presence of increasing urea concentrations. Apparent dissociation constant was 2.04 mu M for the binary complex using solutions of pure beta-LG in agreement with the value for soluble complexes measured by other methods. Hydrophobic interaction appears to be the major force stabilizing complex formation, even though the affinities of the individual caseins do not follow the order of their overall hydrophobicities. Tight binding of micelles to the immobilized sucrose esters suggests that the alkyl chains are able to insert into hydrophobic regions of surface submicelles.