纳豆中抗氧化肽的分离纯化与活性研究

Purification of Antioxidant Peptides from Natto Supernatant and Study on Its Activity

从发酵的纳豆中提取具有抗氧化活性的多肽,通过分子筛层析和反相高效液相色谱对纳豆上清液进行分离纯化,并采用电喷雾串联质谱进行结构鉴定。结果表明:纳豆发酵后的蛋白(多肽)混合物经Sephadex G-50 凝胶色谱进行分离纯化后得到3个组分(F1、F2和F3),其中组分F3的抗氧化活性最强,总还原力达到(8.4±0.6)mmol/g,显著高于其他组分;组分F3经半制备RP-HPLC液相色谱分离纯化得到4个组分(G1、G2、G3和G4),选取含量最高的G1和G3进行活性测定,测得G1的DPPH 自由基清除率达到32.67%。通过 ESI-MS/MS对G1组分抗氧化肽进行一级序列鉴定,得到含量最高和活性最强的10种抗氧化肽序列,并用化学合成的方法对其活性进行验证,得到肽段SFEWVLEH的活性最强,其DPPH清除率为46.66%±0.96%。

High antioxidant polypeptides were extracted from fermented natto,the supernatant of the natto were isolated and purified by SephadexG-50 and RP HPLC,and their structures were identified by electrospray ionization tandem mass spectrometry.The results showed that 3 components(F1,F2 and F3)from the polypeptide mixture of fermented natto were obtained after the separation and purification of the antioxidant peptide via Sephadex G-50 gel.The antioxidant activity of the component F3 was the strongest,the total reduction power reached(8.4+0.6)mmol/g,higher than that of other components.F3 was separated and purified by semi preparative RP-HPLC,and 4 components(G1,G2,G3,and G4)were obtained.The G1 and G3 in the highest contents were selected for determining DPPH scavenging rate,and G1’s DPPH free radical scavenging rate reached 32.67% The antioxidant peptides in G1 were sequenced and identified by ESI-MS/MS.and the 10 antioxidant peptides with the highest content and the strongest activity were obtained.The activity of peptide SFEWVLEH was the strongest verified by chemical synthesis method.The DPPH scavenging rate was 46.66%±0.96%.