Study On The Physico-chemical Characteristics Of ACE Inhibitory Peptides From Lb.plantarum QS670 And Its Synthetic

Yoghurt fermented with Lb.plantarum QS670 was selected as the raw material in this research.It was separated by consecutive purification methods include ultrafiltration,chromatography on Sephadex gel and RP-HPLC to isolate ACE inhibitory peptides.The amino acid sequence of the purified fraction was identified by NanoLC-MS/MS and verified sources and innovativeness by Uni Prot and AHTPDB.This study synthesized ACE inhibitory peptide by solid phase method and explored its physico-chemical characteristics.Experiment results were as follows:1.Three fractions（kDa<3,3>k Da<10,kDa>10）were got from yoghurt supernatant fermented with Lb.plantarum QS670 after ultrafiltration,fraction of kDa<3showed the higher ACE inhibitory activity with 1.998 mg/mL for IC₅₀;The fraction of kDa<3 after freeze-dried and dissolved separated and purified by Sephadex G-15.The best conditions were that the sample weight was 1.5 mL,the sample concentration was200 mg/mL,the elution flow velocity was 0.8 mL/min after conditions optimized.Four fractions were got from the fraction of kDa<3 after Sephadex G-15 under the best conditions,the fraction of D showed the higher ACE inhibitory activity with 1.628mg/mL for IC₅₀;Four fractions were got after RP-HPLC,the fraction of D₁ showed the higher ACE inhibitory activity with 1.258 mg/mL for IC₅₀.2.The dipeptide amino acid sequence Gly-Ala from the fraction of D₁ was identified by NanoLC-MS/MS.Four possible sources and locations,respectivelyαs1-CN（f177-178）,κ-CN（f20-21）,β-LG（f13-14）and SA（f198-199）,were got after compare between GA and main milk protein sequence from the Uni Prot;GA was obtained from fermented milk for the first time through the AHTPDB checked.3.This study synthesized GA by solid phase method.Through mass spectrometry and purity analysis,the sequence and purity of the synthetic were verified.The ACE inhibitory activity IC50 of GA was 1.217 mg/mL,which had no significant difference with the fraction of D₁;4.GA had the high and stable solubleness which higher than 88.2%and ACE inhibitory activity which had the highest ACE inhibitory activity（66.8%）in p H8 between the pH2¹2;The ACE inhibitory activity of GA higher than 63.5%,showed the good stability of ACE inhibitory activity bewteen 20⁶0℃,but the ACE inhibitory activity was markedly decreased with the increased of temperature after 60℃,which showed the lowest ACE inhibitory activity（48.7%）when the temperature down to 100℃;The ACE inhibitory activity of GA was markedly decreased after GA was reacted with pepsin,trypsin and chymotrypsin successively,but still kept over 49.2%,which showed anti-digestion stability.It proved GA was the substrate type ACE inhibitory peptide;The ACE inhibitory activity of GA didn’t have significant difference after react on 0⁸mmol/L of Na⁺,Ca²⁺,Mg²⁺and K⁺,which showed well metal ions stabilit.