Non-covalent complexes of Chlorella pyrenoidosa antioxidant peptides with chlorogenic acid: structure-activity relationships and potential synergistic cytoprotection

CA modulates structures and properties of CPPH via non-covalent binding. • AOPs reveal key electron/H-donating sites underlying complex reactivity. • SAR of AOPs-CA explains assay-dependent synergy patterns in ABTS/DPPH/FRAP. • Quantum parameters link weak binding to reactivity enhancement. • PLR-CA shows potential synergistic cytoprotection in HepG2 cells. Non-covalent complexation with chlorogenic acid (CA) was investigated to remodel complex-level structure-activity relationships of antioxidant peptides (AOPs) from Chlorella pyrenoidosa protein hydrolysates (CPPH) and to evaluate their potential cytoprotective interaction. CPPH-CA complex formation and conformational/microenvironmental changes were characterized by FTIR, intrinsic fluorescence, UV–vis, and interfacial/colloidal measurements. Antioxidant activity and synergy of CPPH-CA and peptide-CA systems were quantified using ABTS, DPPH, and FRAP assays. Representative AOPs were identified by LC-MS/MS and in silico screening, synthesized, and analyzed by quantum chemical parameters (HOMO/LUMO, ESP, ALIE, Fukui functions and IGMH). CA associated with AOPs mainly through hydrogen bonding and van der Waals forces, inducing charge redistribution and decreased HOMO-LUMO gaps, consistent with enhanced reactivity and synergy. In H 2 O 2 -injured HepG2 cells, PLR-CA exhibited a potential cytoprotective effect under the tested conditions by improving cell viability, reducing lipid peroxidation, and restoring antioxidant enzyme activities.