enalapril and hippuryl-histidyl-leucine

Inhibition of angiotensin converting enzyme(EC 3.4,15.1, ACE) in presence of captopril, lisinopril and enalapril were investigated in kidney, lung and serum of sheep using Hip-His-Leu(HHL) as substrate. The activity in kidney, lung and serum was inhibited at HHL concentration above 5 mM. The inhibitory constants (IC50) ranged between 5.6 nM for serum ACE with lisinopril and 70000 nM for renal ACE with enalapril while Ki ranged from 1.0 nM for serum ACE with lisinopril to 12000 nM for kidney ACE with enalapril. Differences in inhibition observed in different tissues suggest that the inhibitors may block function(s) of ACE to varying degrees in each tissue.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Captopril; Enalapril; In Vitro Techniques; Kidney; Kinetics; Lisinopril; Lung; Oligopeptides; Peptidyl-Dipeptidase A; Sheep

A highly sensitive assay for angiotensin I converting enzyme has been developed by using angiotensin I as a substrate. Angiotensin II generated in the reaction mixture was measured by a newly developed specific radioimmunoassay. To protect against angiotensin II destruction, bestatin, an inhibitor of renin, was also used to inhibit plasma renin activity. The reaction was stopped by adding EDTA and MK-521, inhibitors of angiotensin I converting enzyme. The specificity of the antiserum used for the angiotensin II radioimmunoassay was very high. The cross reactivity with angiotensin I was less than 0.5% and none of the proteolytic enzyme inhibitors crossreacted in the assay. The inhibitory effect of pepstatin on plasma renin activity was very high (more than 80%) under the standard assay conditions employed. Serum angiotensinase activity was completely inhibited by the addition of bestatin. An excellent correlation was obtained between this new method and the spectrophotometric method using a synthetic substrate, Hip-His-Leu. The generation of as little as 12 pM of Angiotensin II can be detected. Such low concentration have not been measurable with the usual spectrophotometric method. This new method will facilitate clinical and experimental studies on this unique enzyme, since very low levels of activity can be determined by this highly sensitive radioimmunoassay for angiotensin II.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Angiotensins; Animals; Cross Reactions; Edetic Acid; Enalapril; Leucine; Lisinopril; Oligopeptides; Pepstatins; Peptidyl-Dipeptidase A; Protease Inhibitors; Rabbits; Radioimmunoassay; Renin; Spectrophotometry; Substrate Specificity