超声预处理对金枪鱼皮胶原ACE抑制肽消化稳定性的影响及消化产物的分离纯化、鉴定

Effects of Ultrasound Pretreatment on the Digestion Stability of ACE Inhibitory Peptide from Tuna Skin Collagen and Separation,Purification,Identification of Digestion Products

以金枪鱼皮为原料,超声预处理后酶解制得高血管紧张素转化酶(Angiotensin converting enzyme,ACE)抑制肽,研究ACE抑制肽的模拟胃肠道消化稳定性,并对消化产物进行分离纯化和鉴定。结果表明,超声处理后不同酶解时间酶解液的ACE抑制活性均有所提高;超声预处理20、40 min后,酶解5 min酶解液的消化产物的ACE抑制活性显著高于其他组(p <0.05)。采用葡聚糖凝胶G-15对E5U20和E5U40进行分离纯化,两组酶解液均可分离为3个组分,其中E5U20组组分2的IC_(50)值最小为0.393 mg/mL。采用半制备高效液相色谱对E5U20组分2进行分离纯化,得到8个组分,其中峰3的IC_(50)值最小为0.102 mg/mL。通过质谱法对峰3进行鉴定,测得峰3序列为GPSGPPGP,来源于α1肽链第842～849的位置,符合高ACE抑制活性的多肽构效特点,目前尚无该氨基酸序列的报道。

In this study,tuna skin was prepared by enzymatic hydrolysis after ultrasonic pretreatment to produce hydrolysate with ACE inhibitory activity.This hydrolysate was used as raw material,the gastrointestinal digestion in vitro of hydrolysate was tested.The results showed that all peptides from tuna skin keep high ACE inhibitory activity after digested. After 20 min and 40 min of ultrasonic pretreatment,the ACE inhibitory activity of digested products by enzymatic hydrolysis for 5 min was significantly higher than that of other groups（ p〈0.05）.E5U20 and E5U40 was separated into three components by Sephadex G-15,the peak F2 of E5U20 digestible products was best,of which IC（50） value was 0.393 mg/mL.And then,the F2 was further separated by semi-high performance liquid chromatographic. There were 8 peaks after separated,peak 3 had a lowest IC（50） of 0.102 mg/mL. Finally,the peak 3 was identified by mass spectrometry,the results showed that peak 3 peptide sequence was GPSGPPGP,this sequence was derived from 842 ～ 849 of collagen α1 chain. And these amino acids were consistent with the structure-activity characteristics of ACE inhibitory peptides,and the sequences were not reported by now.