Separation And Purification Of DPP-4 Inhibitory Peptide From Milk Fermented By Lactic Acid Bacteria And Yeast

Diabetes is one of the diseases with the fastest increase in the number of patients in recent decades,and it is a metabolic disease that seriously endangers human health.Studies have found that small-molecule peptides derived from cow's milk can be used as an inhibitor of dipeptidyl peptidase 4(DPP-4),In order to study the inhibitory mechanism of DPP-4 inhibitory peptide in traditional fermented milk.In this thesis,the lactic acid bacteria and yeast fermented milk from traditionally fermented milk in Inner Mongolia were used to determine the DPP-4 inhibition rate to screen out the strains with high DPP-4 inhibition rate.The lactic acid bacteria were identified by 16 S rDNA and the yeasts were identified by 26 S rDNA,and the fermentation conditions were optimized.The fermented milk was digested by simulated gastrointestinal tract,and the DPP-4inhibitory peptide was separated and purified by ultrafiltration membrane and high performance liquid chromatography.Enzyme kinetics was used to determine the action type of the bovine milk-derived DPP-4 inhibitory peptide,and the DPP-4 The effect of inhibitory peptide on DPP-4 enzyme activity in HT29 cells.The main results are as follows: The enzymatic activity determination system of DPP-4 from rabbit small intestine was established.The L34 strain and Y16 strain with the highest DPP-4 inhibitory activity were selected from 100 strains of lactic acid bacteria and 50 strains of yeast.The DPP-4 inhibitory rates were 23.61±0.71% and 31.83±0.47%,respectively,and the L34 strain was identified as Lactococcus lactis,Y16 strain is Yarrowia lipolytica.After optimization of fermentation conditions,the optimal co-fermentation temperature was 31?,the shaker speed was 183r/min,the fermentation time was 76 h,the inoculum was 4%,the inoculation ratio was 1:1,and the DPP-4inhibition rate was 46.59±0.41%,the DPP-4 inhibition rate of fermented milk after simulated gastrointestinal digestion is 57.13±0.85%.The isolated and purified bovine milk-derived DPP-4 inhibitory peptide may have the amino acid sequence of LPLPA and EPVLGPVR.The isolated and purified bovine milk-derived DPP-4 inhibitory peptide may act in a competitive inhibition mode,and can inhibit the DPP-4 enzyme activity in HT29 cells,and its DPP-4 inhibitory rate is 23.41±0.41%.