Preparation And Purification Of Peptides With ACE Ihibitory Activity From Rice Wine

Rice wine is our national specialty, is useful for health preservation,and have very high medicinal value for cardiovascular disease, but there is little experimental data to support the use of Rice wine. This study take the active peptides of rice wine as object, the research is to find methods on isolation and purification active peptides and on the determination of ACE inhibitory activity .The macroporous resin was used to separate peptides from rice wine. By adsorption and desorption experiments, the macroporous resin DA201-C which have the highest adsorption capacity and desorption ratio, was selected from five macroporous resins. Its optimal purification process parameters are determined as follows: extract solution at a concentration of 3.2 mg/ml, 70% ethanol as eluent, elution flow rate 1 BV/h, and its adsorption capacity at 25℃fit best to the Langmuir isotherm. Under these conditions, the purification fold is 9.8, and the yield is 66.7%.HSCCC and Resource 5RPC reversed phase column chromatography were used for the further Separation and Purification. In order to find a suitable solvent system,some vital parameters of the solvent systems were tested. Finally, MtBE:CH₃CN: H₂O (2:2:3,V/V/V)was found to have best performance, so it was used as the two-phase solvent system in the flollowing study. pH-zone-refining countercurrent chromatography was applied to preparative separation of peptides from rice wine. The two phase solvent system composed of MtBE:CH₃CN: H₂O (2:2:3,V/V/V),16mM TFA in organic phase, 8mM NH₃·H₂O in the aqueous mobile phase. The revolution speed: 900rpm; flow rate of the mobile phase:1.8mL/min; temperature:25℃, five components were separated successfully. One of which is P1, was further Purified by Resource 5RPC reversed phase column chromatography. Components P1-1 and P1-2 were found from Components P1.Amino acid composition analysis showed that Component P1-1consisted of Thr,Ser,Gly,Ile and Phe. Among which , Ile and Phe relate to the activity of ACE inhibitor. The vitro ACE inhibitory ability of inhibitors were detected by the HPLC,and it showed that :the inhibitory activity of component P1 was determined as 49.3% with an IC₅₀ of 0.61mg/mL.