Mechanism of interaction between composite GA/EGCG and heat-denatured whey protein by fluorescence quenching

Objective: This study aimed to develop a new carrier system for food functional factors that can effectively regulate the release characteristics during processing, transportation, and storage. Methods: The interaction mechanism between gallic acid （GA） and epigallocatechin gallate （EGCG）, which were combined at different molar ratios （1∶0, 3∶1, 1∶1, 1∶3, 0∶1）, with heat-denatured whey protein isolate （HWPI） was investigated at various temperatures （298.2, 304.2, 310.2 K） using a fluorescence quenching method. Results: The results showed that compared with GA, EGCG had a stronger affinity for HWPI, and when they coexisted, they inhibited each other's binding to HWPI, resulting in decreased binding constants and reduced affinity. Pre-binding of EGCG to HWPI promoted the binding of low concentrations of GA to HWPI, but inhibited the binding of high concentrations of GA to HWPI, pre-binding of GA to HWPI inhibited further binding of EGCG to HWPI. The primary driving forces for binding GA, EGCG, and HWPI were ionic and hydrophobic forces. Compared with the single polyphenol system, the GA/EGCG （3∶1） system had increased ionic and hydrophobic forces in the HWPI reaction, while the GA/EGCG （1∶1） and GA/EGCG （1∶3） systems mainly used hydrophobic interactions as the driving force in the HWPI system. Conclusion: This study demonstrates a competitive relationship among GA, EGCG, and HWPI. The main interaction forces between GA/EGCG and HWPI depend on the composition ratio of the polyphenol complex. Compared to the binary system, the ternary system of GA/EGCG （3∶1） with HWPI exhibits the highest magnitude of ionic and hydrophobic interactions.