Study On Skin ACE Inhibitory Peptides Preparation And Its ACE Inhibitory Activity

Every year there are12million people die of Hypertension around the worldjn china,The morbidity of highpertension is growing with economic growth year by year.We have nearly100million hypertensive patients in china. Each year2million people die of cardiovascular disease.amount of which hypertension is the main factors of cardiovascular disease.Survey Shows,Between patients with hypertension,People who find blood pressure increase account for25%,People who achieve treat account for12%only,People who can control effectively account for3%,Japanese reasearch antihypertensive peptides in1990s,The prevention and control of hypertension still have a long way to go, antihypertensive peptides derive from food proteins with the characteristics of non-toxic side effects, high safety, no blood pressure effect to normal blood pressure person, and suitable for long time use,achieving all countries’attention,This kind of antihypertensive peptide with huge application has become a big lightspot.Now people get antihypertensive peptide derive from fermented milk,Sunflower seeds meal,almond Portein,rice Bran Protein one after another,But antihypertensive peptide derive from CABASA skin have few report.The CABASA fish, Pangasiidae, Pangasius haniltoa, was widely distributed in Mekong river of Vietnam. Which were Special rare high-grade economic fish, in the growth process, there are three pieces of large accumulation of oil in the abdominal cavity, which accounts for about58%of the weight,This fish has the characteristic of growing fast, big body, high output, predatory wide, easy to raise, fleshy white tender, the taste delicious, rich nutrition, no muscle might, good processing etc.This fish has breed successfully in Yingshan country Hubei province.To the use and processing of this kind of fish mainly focus on fish and meat, fish skin,fish bone With rich nutrition value of the products are mostly thrown away directly or as animal feed.CABASA fish was rich in collagen,and could be considered as a potential collagen source, collagen account for74.83%,Collagen was rich in hydrophobic amino acids such as proline Which was reported in literature that it had effect of Antihypertensive activity.so choosing fish skin as the material of producing antihypertensive peptides. Hypertension is kind of Cardiovascular disease which troubling the global world, the major factor which inInducing heart coronary heart disease.Hypertension is often accompanied by heart, blood vessels, brain and kidney and other organs implement qualitative sex pathological change, It causing a variety of complications such as heart, brain, kidney and eye diseases,It is also one of important risk factors of stroke and Coronary heart disease. This article choose CABASA skin as raw materials, then through the Box-Behnken design of response surface optimization of the test, With ACE inhibitory activity being index, from four species commercial Choosing Protamex protease as the experimental enzyme from four kinds of Commercial protease, preparating high activity Antihypertensive peptides, select ultrafiltration, Glucan gel chromatography, High performance liquid chromatographic method isolating and purifying the CABASA skin ACE inhibitory peptides. finally analysis its amino acid sequence, provide an experimental basis for the synthetic ACE inhibitory peptides. The results are as follows:(1) The protease choice experiment:This experiment select alkaline protease, trypsin, Protamex, neutral protease four kinds of protease hydrolysising defatted CABASA skin, Take ACE inhibition rate as index,select Protamex as the experimental enzyme.(2) The parameters optimization test of skim CABASA skin enzymatic hydrolysisAfter choosing enzyme, we carry out Single factor experiment for solid-to-liquid, Hydrolysis time and enzyme quantity,during the Single factor experiment,we choose Hydrolysis time1h,enzyme quantity200u/g,solid-to-liquid1:7-1:9,Based on this design,we carry out Box-Behnken response surface test,Finally we obtain the optimal enzyme solution technology parameters:enzyme quantity340u/g,Hydrolysis time1.27h,In the hydrolysis conditions, The ACE inhibitory activities is IC50=1.417mg/mL.(3) The purification of CABASA skin ACE inhibitory peptide using Ultrafiltration membraneAfter preliminary filtering, ACE inhibitory peptide is five times diluted, then separated by30KD Ultrafiltration membrane, Remove no-hydrolyzed protein, no-substrate substrate and polysaccharide macromolecular impurities. Ultrafiltrati air pressure is controlled in0.2Mpa,collect molecular weight less than30KD liquid and molecular weight more than30KD liquid many times and lyophilizated, assay IC50,The results show that molecular weight less than30KD liquid has higher ACE inhibitory activity.(4)The Sephadex G-50and RP-HPLC purification of CABASA skin ACE inhibitory peptidesUltrafiltrated components is filtered to two absorption peak with a Sephadex G-50gel column,then two chromatography components are studied with ACE inhibitory activity,the1.362mg/mL,0.699mg/mL respectively.we choose components Ⅱ for RP-HPLC purification. (5)The physical and chemical properties of CABASA skin ACE inhibitory peptides.The change of the solubility of the CABASA antihypertensive peptides is very small as the pH between2and10, displaying a high solubility, the solubility is more than80%. Antihypertensive peptides has a strong ability of Hygroscopicity.Emulsification display poor performance when pH appear5, Emulsification display well in Acid environment and alkaline environment. CABASA skin peptide have good thermal stability in high temperature and Room temperature, it have little change after digestive enzymes hydrolysis.