Research On Modification Of Acaudina Molpadioidea Protein Hydrolysates By Plastein Reaction And Its Effects On ACE Activity

Antihypertensive peptides （AHPs） is the inhibitor of angiotensin convertingenzyme （ACE）, which can control blood pressure through reducing the formation ofangiotensinⅡ.AHP has many advantages such as low side effects and toxicity,significant antihypertensive effect, and rich resource.The main contents of this study were as follows:1. Acaudina molpadioidea protein hydrolysates with ACE inhibitory activitywere prepared by complex enzymes and modified by Plastein reaction. The effects ofdifferent kinds of enzymes and the addition level of enzymes, the concentration ofAcaudina molpadioidea protein hydrolysates, temperature and time on the Plasteinreaction of hydrolysates were studied. Acaudina molpadioidea protein hydrolysateswith different modification extents were prepared and their ACE inhibitory activitieswere determined.The results indicated that the ACE inhibitory activity of Acaudinamolpadioidea protein hydrolysates was improved significantly after modified byPlastein reaction.Modified Acaudina molpadioidea protein hydrolysates had adecrease in free amino groups irregularly and their IC₅₀decreased to0.62mg/mL and0.75mg/mL in1h and4h respectively.The decrease of proline in the reaction systemcould significantly increase the ACE inhibitory activity of Acaudina molpadioideaprotein hydrolysates.2. Take Acaudina molpadioidea protein hydrolysates as material, the conditionsof Plastein reaction of Acaudina molpadioidea protein hydrolysates were optimizedby single factor experiment and response surface method,. The optimum conditionsfor Plastein reaction were as follows: concentrations of substrate40.71%, reactiontemperature47.75℃, and enzyme addition levels2.47kU/g pro. Under suchconditions, the decrease of free amino groups was0.462mmol/g pro in6h. 3. Research on the modification of ACE Inhibitory peptides by plastein reactionderived from Acaudina molpadioidea protein in the presence of exogenous Tyr,Pro,Trp,Phe and Leu. Under the optimal reaction conditions, modified products withdifferent modification degrees were prepared by adjusting reaction time. ACEinhibitory activities and IC₅₀values of these modified products were also analyzed.Results indicated that the presence of exogenous Pro can increase the ACE inhibitoryactivity of Acaudina molpadioidea protein hydrolysates. Modified Acaudinamolpadioidea protein hydrolysates had a decrease in free amino groups irregularlyand their IC₅₀decreased to0.59mg/mL and0.63mg/mL from0.89mg/mL afterreaction1h and4h respectively（p＜0.05）.The variations of free amino acidcomposition in reaction system indicate that the content of proline in the reactionsystem decrease significantly.The content of proline decreased to0.23g/（100mL） and0.2g/（100mL） after reaction1h and4h respectively from the beginning of0.29g/（100mL）. It is indicated that proline may bind to the peptide chain, and the ACEinhibitory activity of Acaudina molpadioidea protein hydrolysates were significantlyincreased.4. Two ACE inhibitory activity peptides was isolated from the products ofAcaudina molpadioidea protein hydrolysates modified by Plastein reaction, using thechromatographic methods including Sephadex G-15gel filtration chromatography andreversed phase high-performance liquid chromatography, the sequences were PNVAand PNLG respectively, which deduced as the products of Plastein reaction. Fourpeptides of PNVA, PNLG, NVA, NLG were synthesised by chemical method and theirACE inhibitory activities were determined. The results indicated that the activity ofPNVA （8.18μM） was significantly higher than the activity of NVA （12.69μM）, theactivity of PNLG （12.16μM） was significantly higher than the activity NLG of （17.45μM）. Thus ACE inhibitory activity of the peptides with proline from the products ofPlastein reaction much higher than the original peptides （p＜0.05）, the occurrence ofexogenous proline in the Plastein reaction system improve its ACE inhibitory activity.