ABSTRACT
The mulberry leaf is a botanical resource that possesses a substantial quantity of protein. In this study, alcalase hydrolysis conditions of mulberry leaf protein were optimized using the response surface method. The results showed that the optimum conditions were as follows: substrate protein concentration was 0.5% (w/v), enzymatic hydrolysis temperature was 53.0 °C, enzymatic hydrolysis time was 4.7 h, enzyme amount was 17,800 U/g, and pH was 10.5. Then mulberry leaf peptides were separated by ultrafiltration according to molecular weight. Peptides (<3 kDa) were screened and subsequently identified using LC-MS/MS after the evaluation of α-glucosidase inhibition across various fractions. Three novel potential bioactive peptides RWPFFAFM (1101.32 Da), AAGRLPGY (803.91 Da), and VVRDFHNA (957.04 Da) with the lowest average docking energy were screened for molecular dynamics simulation to examine their binding stability with enzymes in a 37 °C simulated human environment. Finally, they were prepared by solid phase synthesis for in vitro verification. The former two peptides exhibited better IC50 values (1.299 mM and 1.319 mM, respectively). These results suggest that the α-glucosidase inhibitory peptides from mulberry leaf protein are potential functional foods or drugs for diabetes treatment, but further in vivo studies are needed to identify the bioavailability and toxicity.
