Modification of the furanacryloyl-L-phenylalanylglycylglycine assay for determination of angiotensin-I-converting enzyme inhibitory activity.
Angiotensin-I-converting enzyme (ACE, EC 3.4.15.1) plays a central role in the regulation of blood pressure in man. The objective of this study was to evaluate and modify the furanacryloyl-L-phenylalanylglycylglycine (FAPGG) assay method for quantification of ACE activity. The fixed time conditions developed for assay of ACE activity were as follows: 0.8 mM FAPGG, 175 + or - 10 units l(-1) ACE, incubation at 37 degrees C for 30 min and enzyme inactivation with 100 mM ethylenediaminetetra-acetic acid (EDTA). Hydrolysis of FAPGG to FAP and GG was quantified by measuring the decrease in absorbance at 340 nm. It was shown that increasing the level ACE activity in the assay from 155 to 221 + or - 15 units l(-1) resulted in a corresponding increase in the apparent IC(50) value for Captopril from 9.10 to 39.40 nM. Similar trends in the apparent IC50 values for a whey protein hydrolysate were obtained. The results demonstrate the requirement for carefully controlling ACE activity levels in the assay in order to obtained comparable and reproducible values for the inhibitory potency of ACE inhibitors.
Murray
B A
BA
Department of Life Sciences, University of Limerick, Limerick, Ireland.
Walsh
D J
DJ
FitzGerald
R J
RJ
D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology*; D000818:Animals; D002216:Captopril / Q000494:pharmacology; D006033:Glycylglycine / Q000378:metabolism; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D008894:Milk Proteins / Q000378:metabolism / Q000494:pharmacology; D009842:Oligopeptides / Q000032:analysis / Q000378:metabolism*; D007703:Peptidyl-Dipeptidase A / Q000032:analysis / Q000378:metabolism*; D011817:Rabbits; D012015:Reference Standards; D000067816:Whey Proteins
D000806:Angiotensin-Converting Enzyme Inhibitors; D008894:Milk Proteins; D009842:Oligopeptides; D000067816:Whey Proteins; D006033:Glycylglycine; C038275:2-furanacryloyl-phenylalanyl-glycyl-glycine; D002216:Captopril; D007703:Peptidyl-Dipeptidase A