Analysis of Peptides in Fermented Walnut Meal Contributing to Maintain Healthy Blood Glucose Levels and Underlying Hypoglycemic Mechanism

This study was undertaken in order to select peptides with blood glucose regulatory activity from fermented walnut meal with fungi. Animal feeding trials were conducted for overall evaluation of the crude peptide extract, which was subsequently fractionated by Sephadex G-50 column chromatography and high performance liquid chromatography (HPLC). The resulting fractions were tested for inhibitory activity against α-amylase, α-glucosidase and dipeptidase-Ⅳ. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and bioinformatics were combined to determine shared and differential peptides among fractions Ⅳ, Ⅴ and Ⅵ and obtain the major active peptides, which were then quantified by LC. The proteins from which the active peptides were derived were identified by searching the UniProt online platform, and their interactions with the above three enzymes were analyzed by molecular docking. The results showed that the crude peptides effectively controlled body mass, food intake and blood glucose, mitigated insulin resistance and improved glucose tolerance in type Ⅱ diabetic mice. Fractions Ⅳ, Ⅴ and Ⅵ had low molecular mass, which were separated by Sephadex G-50 column chromatography. Fraction V showed no significant difference in inhibitory activity against α-amylase and α-glucosidase but exhibited a significant difference in dipeptidase-Ⅳ inhibitory activity compared with fractions Ⅳ and Ⅵ. Comparative analysis showed that peptides MMRPDEDEQEGQRQ and LPLLR, which were found both in both Ⅴ and Ⅵ, accounted for 82.3% of the total peptide content, and the peptides contributed to 94.3% of the inhibitory activity against DPP-Ⅳ. Therefore, these two peptides are very promising for glycaemic regulation.