Preparation And Separation ACE Inhibitor From Rape Bee Pollen

Angiotensin-converting enzyme (ACE) converts angiotensin I to angiotensinⅡ, and catalyses the degradation of bradykinin, leading to high blood pressure. It can lower blood pressure by inhibiting the angiotensin converting enzyme activity, A number of ACE inhibitory activity hydrolysates have been separated from plant protein.PH stat method, TNBS method and OPA method were compared in determining the degree of rape bee pollen protein hydrolysates, and the pH stat method was selected to detect the degree of rape bee pollen protein hydrolysates, When rape bee pollen protein was hydrolyzed for 0.5mg/ml rape bee pollen protein ACE inhibition rate reached 55.24%, the degree of hydrolysates was 8.97% .Rape bee pollen was hydrolyzed by acid protease, papain, neutral protease and alkaline protease and ACE inhibitory activity was effected by hydrolysates, the corresponding ACE inhibitory activity (IC50) was 0.67mg/mL, 0.55mg/mL, 0.46mg/mL, 0.35mg/mL.Amino acid analysis showed that the concentration of the hydrophobic amino acids from rape bee pollen and hydrolysates in total amino acids was 45.02% and 49.43%, the concentration of alanine was 6.06% and 6.73%, and the concentration of proline was 5.12% and 13.12% respectively.Rape bee pollen hydrolysates was separated by membrane filtration, When the concentration was 0.5mg/mL, the ACE inhibition rate (54.83%) and yield (57%) of the molecular weight bwteen 1500Da and 5000Da hydrolysate was the highest, ACE inhibition rate and yield of greater than 5000Da hydrolysate was 49.64% and 5%; ACE inhibitory rate and yield of bwteen 300Da and 1500Da hydrolysate was 42.72% and 36.55%; ACE inhibition rate and yield of less than 300Da hydrolysate was 5.78% and 1.5%.Four protease hydrolysates were purified by gel and it was found that the components with strong ACE inhibitory activity concentrated between 70min and 120min, and the corresponding molecular weights concentrated between 146Da and 1355Da. ACE inhibition rate of the separate hydrolysates of alkaline protease was the highest, which reached 90%.The six gel components with ACE inhibitory activity were separated from rape bee pollen protein hydrolysate. ACE inhibitory activity of P-1, P-2 and P-3 increased compared with the hydrolysates, the yield was 80.6%; the component with the strongest ACE inhibitory activity was P-2, ACE inhibitory activity (IC50) was 90μg/mL; followed by P-3, ACE inhibitory activity (IC50) was 120μg/mL; the ACE inhibitory activity of P-4, P-5 and P-6 components decreased compared with the hydrolysates, the yield was 17.1%.P-2 was separated by preparative high performance liquid chromatography. ACE inhibitory activity of the separate components was different, the strongest ACE inhibitory activity remains with H-5, IC50 was 10μg/mL; followed by H-3, IC50 was 30μg/mL; the ACE inhibition rates of H-2, H-4 and H-6 were 24.39%, 14.08% and 33.17%; H-1 and H-7 did not have ACE inhibitory activity. H-3 and H-5 need further research, because of their stronger ACE inhibitory activity.When rape bee pollen protein hydrolysates were heated for 20 minutes, ACE inhibition rate of hydrolysates was 49%, with 3.4% being decreased. When rape bee pollen protein hydrolysates were digested by pepsin for 8 hours, ACE inhibition rate of hydrolysates was 37%, which decreased by 27%, When rape bee pollen protein hydrolysates were digested by trypsin for 8 hours, ACE inhibition rate of hydrolysates was 21% with 58% being decreased.