条斑紫菜蛋白酶解液α-葡萄糖苷酶和DPP-Ⅳ抑制活性的表征及肽成分解析

α-Glucosidase and Dipeptidyl Peptidase Ⅳ Inhibitory Activity and Peptide Composition of Porphyra yezoensis Protein Hydrolysate

在特定可控酶解条件下制备获得条斑紫菜1000 Da以下小分子多肽为主的蛋白酶解液,分析发现其具有对α-葡萄糖苷酶和二肽基肽酶Ⅳ(dipeptidyl peptidase Ⅳ,DPP-Ⅳ)的双酶抑制活性,且具有良好的pH值、温度稳定性和胃肠消化耐受性。将酶解液经超滤、纳滤、凝胶过滤层析分离纯化后,采用超高效液相色谱-电喷雾-四极杆飞行时间质谱对活性成分进行鉴定,解析出2条多肽氨基酸序列为Ala-Pro、Pro-Gly-Gly-Val。按上述序列合成出这2条多肽(纯度99.69%)表征其对2种酶的抑制活性,结果表明:Pro-Gly-Gly-Val为双酶抑制肽,它对α-葡萄糖苷酶的IC50为18.60 mmol/L,对DPP-IV的IC50为17.80 mmol/L;而Ala-Pro只对DPP-IV有抑制作用,其IC50为4.65 mmol/L。

Porphyra yezoensis protein hydrolysate,prepared enzymatically under specific and controlled conditions,was found to be dominated by small peptides with molecular mass less than 1000 Da.It had inhibitory activities against α-glucosidase and dipeptidyl peptidase Ⅳ(DPP-Ⅳ)and was able to tolerate pH,temperature and gastrointestinal digestion.The enzymatic hydrolysate was separated and purified consecutively by ultrafiltration,nanofiltration and gel filtration chromatography.Two bioactive peptides,Ala-Pro and Pro-Gly-Gly-Val,were identified by ultra-high performance liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry(UPLC-ESI-Q-TOF-MS).Peptides corresponding to the sequences with a purity of 99.69%were separately synthesized and their inhibitory activities against the two enzymes were characterized.The results showed that Pro-Gly-Gly-Val was a double-enzyme inhibitory peptide with an IC50 of 18.60 mmol/L forα-glucosidase and of 17.80 mmol/L for DPP-Ⅳ.Ala-Pro had an inhibitory effect only against DPP-Ⅳ with an IC50 of 4.65 mmol/L.