Bovine beta-casein micelles as delivery systems for hydrophobic flavonoids

The milk protein beta-casein (beta-CN) is an intrinsically unstructured amphiphilic protein that self-assembles into micelles. Naringenin is the main hydrophobic flavanone in grapefruit and has several beneficial biological effects: it exhibits, for example, antioxidant, anticancer and anti-inflammatory activity. This paper shows that naringenin can be encapsulated in beta-CN micelles. Fluorescence spectroscopy, molecular docking modelling, dynamic light scattering (DLS), static light scattering (SLS) and isothermal titration calorimetry (ITC) were applied to characterize the effect of naringenin on the protein association behavior and properties of the resulting micelles. Naringenin binds to beta-CN at both pH 7 and pH 2, promotes the formation of micelles with a well-defined size distribution and stabilizes the micelles. It was found that naringenin-containing beta-CN micelles have a lower critical micelle concentration (CMC) and a larger aggregation number (N-agg) compared to pure beta-CN micelles. SLS and multi-angle DLS results suggest considerable differences between the structures of pure beta-CN micelles and naringenin-containing beta-CN micelles. In the presence of naringenin spherical micelles were formed with a relatively loose core (hollow sphere), while the pure beta-CN micelles are smaller and seem to be elliptic. Notably, by uptake of naringenin in the micelles, the concentration of naringenin in aqueous solution could be raised considerably. These findings lead to the conclusion that beta-CN micelles are very promising as effective delivery nano-vehicles for hydrophobic bioactive compounds.