Angiotensin I converting enzyme (ACE) inhibitory peptides derived from alfalfa (Medicago sativa L.) leaf protein and its membrane fractions

Alfalfa leaf proteins (ALP) were extracted through the method of alkali solubilization and acid precipitation, and the six different ALP hydrolysates were obtained by hydrolysis of papain, Alcalase, Neutrase, trypsin, pepsin, and Flavourzyme, respectively. The results showed that alfalfa leaf protein hydrolysate (ALPH) prepared by papain exhibited the highest angiotensin I converting enzyme (ACE) inhibitory activity of 79.69% +/- 2.77% at the same concentration. Hence, papain ultimately was used to hydrolyze ALP for producing ACE inhibitory activity peptides. The hydrolysis conditions of hydrolysis temperature, enzyme to substrate ratio (E/S), and pH were optimized by using response surface methodology (RSM). We obtained the maximum ACE inhibitory rate of 92.33% +/- 0.30% under E/S of 3%, pH of 7.5 and hydrolysis temperature of 55 oC. Finally, the hydrolysate was separated into four ranges of molecular weight (ALPH-1, >30 kDa; ALPH-2, 30-10 kDa; ALPH-3, 10-5 kDa; ALPH-4, <5 kDa) through different ultrafiltration membranes. Among them, ALPH-4 showed the maximum ACE inhibitory activity of 93.85% +/- 1.09%, implying that ALPH-4 should be considered as a potential source of natural ACE inhibitor. Novelty impact statement: Recently, more and more researches focus on ACE inhibitors which are derived from food because they are safer than the synthetic chemicals which may result in several adverse events. We found that ACE inhibitory peptides from Alfalfa leaf protein have the high ACE inhibitory activity in vitro, which could be considered as a potential approach for use in nutraceuticals for the prevention and management of hypertension. People have a high demand to consume functional products, this has driven the food industry to the development of a new commercial ACE inhibitory peptides products in the market.

Automated summary

Alfalfa leaf proteins were hydrolyzed using different enzymes, with papain showing the highest ACE inhibitory activity. Optimal hydrolysis conditions were determined through response surface methodology, resulting in a high ACE inhibitory rate. The hydrolysate was separated based on molecular weight, with ALPH-4 exhibiting the highest ACE inhibitory activity, suggesting it as a potential natural ACE inhibitor.