| Whey protein hydrolysis(WPH)and its bioactive peptides have a variety of biological activities,especially their effects on regulating blood sugar have been widely concerned,with profound research value.Whey protein is mainly composed of beta-lactoglobulin and alphalactalbumin,which also contains high concentration of branched-chain amino acids,is an excellent source of bioactive peptides.Recent studies have shown that the active peptides in WPH can inhibit Dipeptidyl peptidase-IV(DPP-Ⅳ)activity,promote glucagon-like peptide secretion,improve blood sugar regulation and inhibit food intake.In this study,whey protein was hydrolyzed by human digestion of common pepsin and trypsin,and the hydrolysis conditions were optimized.The effects of various factors on the hydrolysis degree and DPP-Ⅳ inhibition rate of WPH were determined by single factor test,and the hydrolysis conditions were optimized by response surface methodology.The changes of free amino acid content in the hydrolysis process were studied to determine the hydrolysis conditions of WPH.The digestive characteristics were evaluated,and the inhibitory activity of DPP-Ⅳ of Momordica charantia saponins(MS)was studied.The synergistic effects of DPP-Ⅳ of MS and hydrolysates of whey protein were studied.The changes of DPP-Ⅳ inhibitory activity of hydrolysates and MS after digestion were determined to evaluate the digestive characteristics.The main conclusions are as follows:(1)Both WPH of pepsin and trypsin have DPP-Ⅳ inhibitory activity.The hydrolysis degree and DPP-Ⅳ inhibition rate of whey protein pepsin hydrolysate were different under different hydrolysis time and enzyme-substrate ratio.The optimum hydrolysis conditions were determined by single factor experiments as follows: hydrolysis time 2 h,hydrolysis p H 2.5,pepsin to substrate ratio 4%.Response surface analysis showed that the optimum hydrolysis conditions were p H2.69,hydrolysis time 2.05 h and the ratio of enzyme to substrate 4%.The IC50 value of whey protein pepsin hydrolysates for DPP-Ⅳ inhibitory under this condition was calculated to be 5.77 mg/m L.The hydrolysis degree and DPP-Ⅳ inhibition rate of WPH of trypsin were different under different hydrolysis time and enzyme-substrate ratio.The optimum hydrolysis conditions were determined by single factor experiments,were 4 h,p H 8 and 4%.Response surface analysis showed that the optimum hydrolysis conditions were 3.99 h,p H 7.92 and 4%.The IC50 value of whey protein pepsin hydrolysates for DPP-Ⅳ inhibitory under this condition was calculated to be 7.28 mg/m L.(2)Gastrointestinal digestion simulation test was carried out on the two hydrolysates,and the content of free amino acids in the hydrolysates was determined.The total content of free amino acids in pepsin hydrolysates was 47.00 mg/m L.After simulated gastrointestinal digestion and intestinal digestion,it increased to 60.08 mg/g and 57.99 mg/g,respectively.And the content of leucine was higher than that of free amino acids in tryptic hydrolysates.While the WPH of tryptin after simulated gastrointestinal digestion and intestinal digestion,it increase from 42.95 mg/g to 58.12 mg/g and 52.87 mg/g respectively,and the content of tyrosine was significantly higher than other amino acids(P<0.05).(3)MS have DPP-Ⅳ inhibitory activity.The IC50 value of Momordica charantia saponins is 5.20 mg/m L.When the ratio of MS to hydrolysate is 1:1,the inhibitory effect of the mixture is significantly higher than the sum of these alone(P<0.05).The DPP-Ⅳ inhibition rates of WPH of pepsin and trypsin and MS were all changed after simulated gastrointestinal digestion.The inhibition rates of WPH of pepsin after simulated gastrointestinal digestion were significantly reduced(P<0.05),while the inhibition rates after simulated gastrointestinal digestion were significantly increased(P<0.05);the inhibition rate of WPH of trypsin after simulated gastrointestinal digestion increased significantly(P<0.05),but decreased significantly after simulated gastrointestinal digestion(P<0.05);the inhibition rate of MS after simulated gastrointestinal digestion increased significantly(P<0.05),but decreased significantly after simulated gastrointestinal digestion(P<0.05).In conclusion,in vitro experiments showed that both WPHs and Momordica charantia saponins had inhibitory activities against DPP-Ⅳ.The combination of hydrolysis conditions with the highest inhibitory rate of pepsin and whey protease hydrolysate DPP-Ⅳ was determined.The IC50 values of the two hydrolysates were calculated.The simulated gastrointestinal tract test was carried out.The content of free amino acids and DPP-Ⅳ inhibitory rate in hydrolysate changed before and after digestion.The IC50 value of MS is calculated,which provides a reference for the study of the mechanism of regulating blood sugar by MS,and is of great significance for the further processing of whey and Momordica charantia and its application in regulating blood sugar activity. |
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