Abstract: The interaction and mechanism of casein (CA) with hesperidin (HES) and naringin (NAR) were investigated using fluorescence spectroscopy, synchronous fluorescence spectroscopy, thermodynamic analysis, and molecular docking in this study. The results showed that both polyphenols quenched CA in a static manner, but HES had stronger fluorescence quenching effect and binding affinity to CA and had more binding sites. According to thermodynamic parameters, the binding of CA to HES and NAR was spontaneous and mainly driven by hydrophobic forces. According to synchronous fluorescence spectra, introduction of the two polyphenols caused changes in the hydrophobic environment of amino acid residues in CA. The binding process was mostly driven by hydrophobic forces according to molecular docking, and more hydrogen bonds were formed between HES and CA, making the binding between them more stable. The results of this study can be used as a reference for future studies on polyphenol-protein interactions as well as the feasibility of CA as a carrier of flavonoid polyphenols.

Key words: fluorescence spectroscopy; hesperidin; naringin; casein; molecular docking