A rapid assay for angiotensin-converting enzyme activity using ultra-performance liquid chromatography-mass...

Angiotensin-converting enzyme (ACE) plays an important role in the renin-angiotensin system and ACE activity is usually assayed in vitro by monitoring the transformation from a substrate to the product catalyzed by ACE. A rapid and sensitive analysis method or ACE activity by quantifying simultaneously the substrate hippuryl-histidyl-leucine and its product hippuric acid using an ultra-performance liquid chromatography coupled with electrospray ionization-mass spectrometry (UPLC-MS) was first developed and applied to assay the inhibitory activities against ACE of several natural phenolic compounds. The established UPLC-MS method showed obvious advantages over the conventional HPLC analysis in shortened running time (3.5 min), lower limit of detection (5 pg) and limit of quantification (18 pg), and high selectivity aided by MS detection in selected ion monitoring (SIM) mode. Among the six natural products screened, five compounds, caffeic acid, caffeoyl acetate, ferulic acid, chlorogenic acid and resveratrol indicated potent in vitro ACE inhibitory activity with IC(50) values of 2.527 +/- 0.032, 3.129 +/- 0.016, 10.898 +/- 0.430, 15.076 +/- 1.211 and 6.359 +/- 0.086 mm, respectively. A structure-activity relationship estimation suggested that the number and the situation of the hydroxyls on the benzene rings and the acrylic acid groups may play the most predominant role in their ACE inhibitory activity.