Kinase experiment:

Hip-His-Leu (5mM) and an ACE inhibitor (Leucylarginylproline) are dissolved in a 100 mM sodium borate buffer (pH 8.3) containing 300 mM NaCl, and incubated for 30 min with 8 milliunits of ACE at 37℃. The ACE inhibitor concentration required to inhibit 50% of the ACE activity under the above conditions is defined as IC50[1].

Animal experiment:

Rats: Leucylarginylproline is dissolved in 1.0 mM normal saline. After being warmed up, rats are orally administered (0.18 mmol/kg bw) with peptides or normal saline (control). Tail systolic blood pressures are measured at 2-h intervals (0, 2, 4, 6, 8, 10, and 12 h) after the administration[2].

Leucylarginylproline is an angiotensin-converting enzyme (ACE) inhibitor with an IC50 of 0.27μM.

Intravenous injection of Leucylarginylproline (30mg/kg) causes a decrease in the blood pressure. The maximum mean blood pressure reduction (about 15 mmHg) occurrs about 2 min after the injection[1]. Leucylarginylproline peptide reduces the blood pressure by about 15 mmHg at the fourth hour and shows a maximal reduction effect of about 35 mmHg at the eighth hour after oral administration[2].

[1]. Miyoshi S, et al. Structures and activity of angiotensin-converting enzyme inhibitors in an alpha-zein hydrolysate. Agric Biol Chem. 1991 May;55(5):1313-8. [2]. Chen TL, et al. Microencapsulation and modification of synthetic peptides of food proteins reduces the blood pressure of spontaneously hypertensive rats. J Agric Food Chem. 2003 Mar 12;51(6):1671-5.