Enhanced phloretin binding and bioactivity by ultrasonication-induced structural modulation of ovalbumin: insights...

The practical utility of phloretin (PHL), a dihydrochalcone flavonoid, is significantly hampered by its low solubility and bioavailability. In this study, ovalbumin (OVA) was used as a carrier to prepare UOVA-PHL (ultrasonication before binding) and U(OVA-PHL) (binding before ultrasonication) at ultrasonication durations of 30, 50, and 70 min, with non-ultrasonicated OVA-PHL as the control. The effects of the ultrasonication sequence on the interactions within the complexes and PHL bioactivity were evaluated. Ultrasonication increased the PHL binding content of OVA, with the highest value observed at 50 min. At this ultrasonication duration, the most pronounced changes in the OVA secondary structure, hydrodynamic diameter, and ζ-potential were also detected. Fluorescence spectroscopy indicated hydrogen bonding and van der Waals forces as the primary intermolecular forces, with binding affinity in the order UOVA-PHL > U(OVA-PHL) > OVA-PHL and surface hydrophobicity in the reverse order. Molecular dynamics simulations were performed to analyze conformational dynamics, solvation, hydrogen bonding, and binding free energy. The results further confirmed the ultrasonication-induced structural modulation and the strongest binding stability of UOVA-PHL. Furthermore, evaluations of stability, solubility at different pH levels, simulated gastrointestinal digestion, extracellular and intracellular antioxidant activities, and cytotoxicity demonstrated that OVA encapsulation and ultrasonication improved the functionality of PHL. Ultrasonication before binding effectively unfolded OVA, enhanced affinity, and significantly improved PHL bioaccessibility and antioxidant activity. This study provides a rationale for designing ultrasound-assisted protein-polyphenol nanocarriers and lays an experimental foundation for enhancing the application potential of PHL in functional foods.