Identification of binding sites for Tartary buckwheat protein-phenols covalent complex and alterations in protein structure and antioxidant properties.

To investigate the effects of structure, multiple binding sites and antioxidant property of Tartary buckwheat protein-phenols covalent complex, protein was combined with different concentrations of phenolic extract. Four kinds of phenols were identified by UPLC-Q/TOF-MS, which were rutin, quercetin, kaempferol and myricetin. UV-vis absorption spectroscopy and X-ray diffraction showed that the phenols can successfully bind to BPI. Fourier-transform infrared, circular dichroism and fluorescence emission spectroscopy showed that the binding of phenol can change the secondary/tertiary structure of protein. The particle distribution indicated that the binding of phenols could reduce the particle size (from 304.70 to 205.55 nm), but cross-linking occurred (435.35 nm) when the bound phenol content was too high. Proteomics showed that only rutin, quercetin and myricetin can covalently bind to BPI. Meanwhile, 4 peptides covalently bound to phenols were identified. The DPPH· scavenging capacity of complexes were from 8.38 to 33.76 %, and the ABTS·+ binding activity of complexes were from 19.35 to 63.99 %. The antioxidant activity of the complex was significantly higher than that of the pure protein. These results indicated that protein-phenol covalent complexes had great potential as functional components in the food field.