Abstract: The objective of the present study was to purify and structurally identify hypoglycemic peptides from wheat germ protein. The protein was hydrolyzed with trypsin to obtain the hypoglycemic peptides, and the hypoglycemic effect was evaluated by animal test. The active peptides were obtained by ultrafiltration. The optimum resin was selected for purification of the peptides by ion exchange adsorption, and further purification was carried out by sequential column chromatography on SephadexG-25 and SephadexG-15 before separation by reverse phase-high performance liquid chromatography (RP-HPLC). Finally, the structures of the purified peptides were identified by HPLC-electrospray ionization-mass spectrometry (HPLC-ESI-MS). The results showed that the enzymatic hydrolysate had α-glucosidase inhibitory activity with an IC50 of 10.98 mg/mL, and could relieve symptoms in diabetic mice. The highly active peptides with molecular mass less than 5 kDa were obtained by ultrafiltration, and their IC50 was 1.60 mg/mL. 732 cation exchange resin showed the best separation efficiency, and the elution percentage with 2.5% ammonia was 98.13%. After ion exchange adsorption, the hypoglycemic activity was significantly improved, yielding an IC50 of 0.30 mg/mL. Eight peaks were obtained after purification by RP-HPLC and among these the amount and activity of peak 1, with an IC50 of 0.098 mg/mL, were both highest. HPLC-ESI-MS analysis showed that the abundance at m/z 274.45 was high and that 7 peptides including 1 dipeptide and 6 tripeptides were obtained by ion fragment re-assembling. The findings obtained in this work are of significant importance for the development of novel functional foods with hypoglycemic efficacy.

Key words: wheat germ, hypoglycemic peptides, α-glucosidase, isolation and purification, structural identification, animal test