Delicious Leather Pouch Siphon Worm Protein Extraction And Enzyme Legal Alternate Research Of Antihypertensive Peptides

Phascolosoma esculenta is one of marine creatures, rich in protein, low in fat, with18kinds of amino acids. Angiotensin Ⅰ-converting enzyme (ACE) plays an important part in blood pressure regulations. In this work, preparation of ACEI (inhibitor of ACE) from Phascolosoma esculenta protein was studied. The work could be divided into four sections.The first section, extraction of Phascolosoma esculenta protein was researched; the optimal process is1%(w/v) NaCl,1:20(solid-liquid),50℃,4h, with the ratio up to42%. SDS-PAGE shows that molecular weight of protein mainly distributed in110,80,44,36,18,12kDa. The activity of protein in inhibiting ACE was low.The second section, Phascolosoma esculenta water-soluble protein hydrolysed by pepsin, trypsin, neutral protease, alkali protease and papain were researched. Pepsin is the best enzyme in hydrolying protein, with the highest ratio of inhibiting the ACE. The IC50of hydrolysates is1.85mg/ml. The IC50value was defined as the concentration of inhibitor that could inhibit50%of the ACE activity.The third section, purification, structure identification and activity of hydrolysates were researched, conclusions were as follows.1) Ultrafiltration, ion exchange, sephadex and RP-HPLC technologies were used in purification of hydrolysates. The value of IC50increased along with purification, from1.85mg/ml of hydrolysates to0.165mg/ml of component p1-d-Ⅲ(P), with10times higher.2) Analysis of P by MALDI-TOF/TOF was also researched, the Mw and amino acid sequence of P were1222.7Da and Ala-Trp-Leu-His-Pro-Gly-Ala-Pro-Lys-Val-Phe, which is a novel antihypertensive peptides, named XFpI.3) Properties of P was examined, ACE-inhibitory activity showed little change after incubation in different temperature and pH and with simulated gastric juice, which suggests that these peptides might be resistant to digestion in the gastrointestinal tract.4) ACE inhibition pattern of the purified ACE inhibitor from enzymolysis of Phascolosoma esculenta was investigated by Lineweave-Burk plot. It was found to be a competitive inhibition pattern on ACE.The forth section, The activity of ACE inhibitor in vivo was researched with spontaneously hypertensive rats (SHR) and result showed that blood pressure start to decrease significantly at2h after oral administration. Compared with the sipunculid protein, peptic hydrolysate and peptide P show higher activity in reducing the blood pressure, SBP reduced24and43mmHg, especially peptide, with higher activity and maintaining longer hours.