Oxidases and rolling dominate black tea polysaccharide conjugates formation: Enzymatic and non-enzymatic mechanisms

Tea processing significantly contributes to the heterogeneity of tea polysaccharide conjugates; however, the specific mechanisms underlying their formation remain unclear. In the present work, it is found that rolling is the critical stage for the formation of black tea polysaccharide conjugates. In vitro enzymatic reactions demonstrated that oxidases, particularly peroxidase with synergistic support from polyphenol oxidase, mediate conjugate formation. Molecular docking studies further revealed strong interactions between proteins, phenolics, and sugar chain units through hydrogen bonding and hydrophobic forces, with key residues such as LEU-189, ARG-458, and HIS-145 playing pivotal roles. This study elucidates the enzymatic and non-enzymatic mechanisms underlying black tea polysaccharide conjugate formation and provides novel insights for optimizing tea processing to precisely design polysaccharide conjugates.