小米糠多肽抗氧化活性及对秀丽隐杆线虫延寿作用初析

Antioxidant Activity of Polypeptides from Millet Bran and Its Lifespan-Prolonging Effect on Caenorhabditis elegans

为了提高小米糠的利用率,为小米糠的综合利用与高值开发奠定一定理论基础,采用碱溶酸沉法制备了小米糠蛋白,用碱性蛋白酶酶解后使用超滤管对酶解的蛋白样品进行超滤,从而得到3组不同分子质量(MW)的多肽组分(MW<3 ku、3 ku<MW<10 ku和MW>10 ku),进一步对这3种组分小米糠多肽的抗氧化活性进行研究。结果表明,MW<3 ku的多肽组分体外抗氧化活性最佳,当多肽质量浓度为6 mg/mL时,对ABTS自由基、DPPH自由基和羟自由基的清除率分别达到了90.28%、78.43%和34.23%,具有较强的自由基清除能力。将MW<3 ku的小米糠多肽组分作用于秀丽隐杆线虫后,线虫寿命较对照组显著提高,在质量浓度为20μg/mL时延长寿命效果最佳;线虫运动和咽抽能力试验结果表明,20μg/mL的小米糠多肽能显著增强线虫运动能力和咽抽速率。进一步采用液质联用技术(LC-MS/MS)对小米糠多肽进行了测序,并结合分子对接技术预测了小米糠代表性肽ASEGGHGPHWPLPPF与髓过氧化物酶(Myeloperoxidase,MPO)的作用方式,结果显示,该肽与MPO活性位点具有强烈的分子相互作用,推测其具有抵抗体内氧化应激的能力。

In order to improve the utilization of millet bran,lay a foundation for the comprehensive utilization and the preparation of high add-value products of millet bran,in this study,millet bran protein was extracted by alkali-solution and acidisolation.The hydrolyzations of protein samples by alcalase were ultra-filtrated by hyperfiltration tube to obtain three polypeptide components with different molecular weights(MW<3 ku,3 ku<MW<10 ku,and MW>10 ku).The antioxidant activity of the three components of millet bran polypeptide(MBPE)was further studied.The results showed that the peptide component with the molecular weight of less than 3 ku exhibited the strongest antioxidant activity.When the mass concentration of polypeptide component was 6 mg/mL,its scavenging rate of ABTS radical,DPPH radical,and hydroxyl radical reached 90.28%,78.43%,and 34.23%,respectively,showing strong scavenging ability of free radicals.The lifespan of C.elegans with the polypeptide component with MW<3 ku significantly increased compared with the control and the best lifespanprolonging effect occurred at the mass concentration of 20μg/mL.The exercise and pharyngeal pumping ability of C.elegans tests showed that 20μg/mL of millet bran polypeptides could significantly enhance exercise ability and pharyngeal pumping rate of C.elegans.The millet bran polypeptides were sequenced by liquid chromatography-mass spectrometry(LC-MS/MS).Combined with the molecular docking technique,action mode of ASEGGHGPHWPLPPF which was the representative peptide sequence from miller bran and human myeloperoxidase(MPO)was predicted,and the results indicated that the peptide had a strong molecular interaction with the active site of myeloperoxidase,which was speculated to have the ability to resist oxidative stress in vivo.