Complex enzymolysis-derived peanut peptides: in silico selection of antioxidant peptides, emulsion-microcapsule encapsulation, and in vitro digestive performance evaluation.
Peanut peptides (PP) are promising plant-derived functional ingredients, but bitterness and gastrointestinal instability limit oral applications. This study integrated complex enzymolysis, LC-MS/MS-assisted in silico screening, molecular dynamics (MD), and W1/O/W2 double-emulsion microencapsulation to establish a laboratory-scale discovery-to-delivery workflow for antioxidant PP. Under optimized alkaline protease/flavor protease hydrolysis (5:5, w/w; 10% enzyme; 59 °C; 9 h), LC-MS/MS identified 763 peanut-derived peptides. Stepwise screening using PeptideRanker (>0.5), ToxinPred3.0 (<0.5), and AlgPred2.0 (<0.5) yielded 157 candidates. CDOCKER docking identified P152 (FEELNADLFR) and P142 (YFPTQALNFAFK) as the top docking-ranked Keap1-Kelch and MPO candidates, respectively, with CDOCKER energies of -163.791 and - 158.855 kcal/mol; P34 (SPNVDPPKTP) and P135 (IPVPFPDPDGDYT) showed the lowest calculated binding energies with ABTS+ and DPPH, respectively. MD/MM-PBSA further supported dynamic stability and favorable binding tendencies of P152-Keap1-Kelch and P142-MPO complexes. The optimized PP-loaded double-emulsion microcapsule (PP-M) showed high apparent encapsulation efficiency (89.43%, nitrogen-balance estimate). Human sensory evaluation and electronic-tongue analysis demonstrated that PP-M significantly reduced perceived bitterness and instrumental bitterness response compared with free PP hydrolysate at matched peptide-equivalent concentration. Static INFOGEST digestion indicated gastric structural retention followed by intestinal disintegration, accompanied by increased peptide-equivalent content and enhanced radical-scavenging activity in the digesta. Overall, PP-M improved the in vitro sensory acceptability and bioaccessibility of PP, supporting formulation-level bitterness alleviation rather than complete bitterness elimination. Targeted bitter-peptide identification, marker-peptide release profiling, epithelial transport, and in vivo validation remain necessary.