<p>Bioactive peptides demonstrate multiple biological effects which advance health benefits, particularly antioxidant capacities, antimicrobial effects and immunomodulatory activities. A range of conventional methods has been applied to obtain peptide chains such as enzymatic hydrolyzation and solvent extraction; however, it may disturb peptide properties through drastic structural changes. This review suggests synergistic methods of novel technologies followed by enzymatic hydrolyzation yield peptide fractions with high purity and maintained bioactive qualities. The peptide configuration is emphasized as it constitutes a fundamental factor towards functional properties including amino acid compositions and their arrangement, molecular size and secondary structural alteration. This further elucidates comprehensive insights into the underlying mechanisms of biological activities associated with peptide conformation, molecular interaction and signaling pathways. The engagement of peptides with many molecular types offers alternative approaches in enhancing free radical scavenging and microbial inhibition characteristics. Furthermore, the current study consolidates the application of peptide ingredients across various food products, addressing underexplored perspectives and advancing industrial uses with the increased peptide stability. The integration of computational bioinformatic tools and artificial intelligence (AI) is revealed to fulfill the challenges of peptide fractionation, forecasting the structure–activity relationship precisely.</p> Graphical Abstract <p></p>