金枪鱼(Katsuwonus pelamis)碎肉蛋白降压肽的酶解制备及活性研究

以血管紧张素转换酶(ACE)抑制率为指标,通过正交试验L9(34)确定碱性蛋白酶(Alcalase)对金枪鱼(Katsuwonus pelamis)碎肉蛋白的最佳酶解条件,利用D101大孔树脂建立了酶解物的脱盐工艺,利用超滤、葡聚糖凝胶(G-25)色谱和反相-高效液相色谱(RP-HPLC)分离降压肽,并确定其纯度,利用氨基酸序列分析和ESI-MS鉴定纯化多肽的结构。结果表明,碱性蛋白酶酶解金枪鱼碎肉蛋白制备降压肽的最佳酶解条件是:pH 9.5,酶用量1.5%,酶解温度50°C,酶解时间5h;D101大孔吸附树脂脱盐工艺条件为:上样浓度10 mg/mL、上样流速1.5 BV/h、解吸剂为75%乙醇;酶解物经超滤和Sephadex G-25分离获得一个纯度较高的四肽,经氨基酸序列分析和ESI-MS鉴定结构为Phe-GlyGly-Val(FGGV),ESI-MS检测给出分子离子峰m/z 379.50(M+H]+)。利用碱性蛋白酶酶解并经超滤和色谱技术制备的金枪鱼碎肉蛋白降压肽具有良好的ACE抑制活性,可作为降压药物、保健食品或添加剂进行开发。

PREPARATION AND CHARACTERISATION OF AN ANTIHYPERTENSIVE PEPTIDE FROM PROTEIN HYDROLYSATE OF ASKIPJACK TUNA (KATSUWONUS PELAMIS) GROUND MEAT

In this text, the conditions of alcalase hydrolysis on askipjack tuna (Katsuwonus pelamis) ground meat and desalting process of the protein hydrolysate were optimized, and the antihypertensive peptide was isolated from the hydrolysate by using ultrafiltration and gel filtration chromatography, and its structure was determined using ESI-MS and Procise Protein/Peptide Sequencer. The protein hydrolysate with high ACE inhibition ratio was gained under the optimal conditions of temperature 50?C, pH 9.5, enzymatic hydrolysis time 5 h, and amount of enzyme 1.5%. The condition for desalting process was optimized as followed: loading sample concentration of 10 mg/mL, sampling flow rate of 1.5 BV/h and 75% alcohol as eluent. The fractions and purified peptide from desalting hydrolysate were prepared by using ultrafiltration and Sephadex G-25 column, and a tetrapeptide with high ACE inhibition was isolated and identified as Phe-Gly-Gly-Val (FGGV). The present results suggested that the protein hydrolysate, fractions and purified peptide of tuna ground meat could be used as natural antihypertensive drugs, foods or additives.