低值紫菜蛋白酶解产物中抗氧化活性肽的纯化及分析鉴定

Purification and Identification of Antioxidant Peptides from the Product of Protease Hydrolysis the Low-Cost Laver

采用中性蛋白酶和氨肽酶复配酶解低值紫菜,体积分数60%乙醇沉淀去多糖,上清液为紫菜抗氧化多肽粗品,该粗品利用Sephadex G-10、DEAE-52和SOURCE 3RPC等色谱手段分离纯化,得到酶解紫菜抗氧化活性肽(ELAP)。当ELAP的质量浓度为100μg/mL时,其对超氧自由基、DPPH和羟自由基的清除率依次为23.25%,47.12%,47.12%,分别是同浓度下抗坏血酸(VC)的0.79,0.87,0.91倍,还原能力为VC的1.23倍,有很强的抗氧化性。ELAP经反相高效液相(RP-HPLC)分析型色谱测定其纯度达87.44%。超高效液相色谱-质谱法(Q-TOFMS)分析鉴定ELAP为一种6肽DGVGYG(Asp-Gly-Val-Gly-Tyr-Gly),其组成中Tyr与Gly本身就属于具抗氧化活性的氨基酸。

The low-cost laver was hydrolyzed with neutral protease and aminopeptidase in this study.The antioxidative peptides were purified initially by adding 60%ethanol to remove polysaccharides in the hydrolysate.The supernatant was further purified through Sephadex G-10 gel,DEAE-52 anion-exchange and SOURCE 3RPC to get more purer enzymatic laver antioxidative peptides(ELAP).The scavenging activities of Superoxide-radical,DPPH radical and Hydroxyl -radical were 23.25%,47.12%and 47.12%respectively when the content of the ELAP was at 100μg/mL.Compared to vitamin C at the same content(100μg/mL),the scavenging activities in the ELAP were 0.79,0.87 and 0.91 times of the activities in vitamin C,Furthermore, the reducing ability of the low-cost laver peptides was 1.23 times than that of vitamin C. According to the chromatographic of analytical reverse-phase high performance liquid chromatography (RP-HPLC).the purity of ELAP was up to 87.44%.The ELAP was identified as a hexapeptide AGVGTG(Asp-Gly-Val-Gly-Tyr-Gly) with analysis of the ultra-performance liq- uid chromatography-quadrupole time-of-flight mass spectrometry(Q-TOF-MS).The composition of the hexapeptideC AGVGTG) in the ELAP which contained Tyr and Gly has the typical function of antioxidation.