Reducing, radical scavenging, and chelation properties of in vitro digests of alcalase-treated zein hydrolysate

The objective of the study was to assess the antioxidant potential of alcalase-treated zein hydrolysate (ZH) during a two-stage (1 h of pepsin → 0.5-2 h of pancreatin, 37°C) in vitro digestion. Sephadex gel filtration and high-performance size exclusion chromatography were used to separate ZH into fractions. The amino acid composition, 2,2′-azinobis(3- ethylbenzothiazoline-6-sulfonic acid) (ABTS^+•) and 1,1-diphenyl-2-picrylhydrazyl (DPPH^•) free radical scavenging activity, reducing power, and Cu²⁺ chelation ability were tested to determine the antioxidant efficacy of ZH. Results showed that in vitro digests of ZH contained up to 16.5% free amino acids, with short peptides (<500 Da) making up the rest of the mass. The ABTS^+• scavenging activity of ZH was decreased by 27% (P < 0.05) after pepsin treatment but was fully recovered upon subsequent pancreatin digestion, while the DPPH^• scavenging activity of ZH was substantially less than ABTS^+• scavenging activity and showed a 7-fold reduction following pancreatin treatment. The reducing power of ZH increased 2-fold (P < 0.05) following pancreatin digestion when compared with nondigested ZH. The ability of ZH to sequester Cu²⁺ was reduced by pepsin digestion but was reestablished following pancreatin treatment. The antioxidant activity demonstrated by in vitro digests of ZH (1-8 mg/mL) was comparable to or exceeded (P < 0.05) that of 0.1 mg/mL of ascorbic acid or BHA. The results suggested that dietary zein alcalase hydrolysate may have the benefit to promote the health of the human digestive tract.