Solubilization of genistein by caseinate micellar system

This study investigates the aggregation behavior of caseinate and the solubilization of genistein in aqueous caseinate solution. The critical aggregation concentration (CAC) of caseinate was obtained from the fluorescence intensity of 8-anilino-1-naphthalenesulfonic acid (ANS), which was enhanced by ANS-protein interactions and the hydrophobicity of caseinate. The increasing solubility of genistein in caseinate was confirmed by HPLC measurements; above and below the CAC, the genistein/caseinate molar ratio is 1:1 and 10:1, respectively. The latter ratio indicates that more caseinate molecules surround genistein below the CAC. However, the solubility of genistein in caseinate is unaffected by calcium ions. Atomic force microscopy (AFM) shows that casein sub-micelles are similarly structured in the presence and absence of genistein. In AFM phase images, the caseinate sub-micelle is brightened in the presence of genistein, implying that the particle becomes more rigid, probably because genistein attaches to the surface or to the narrow part of the sub-micelle. The diameter of sub-micelle aggregates is two times that of caseinate alone (24 nm versus 12 nm). These results were confirmed by cryo-TEM observations.