The rate of loss of phenol or 4-nitrophenol from N alpha-(diphenoxyphosphoryl)-L-alanyl-L-proline (2), N alpha-[bis(4-nitrophenoxy)phosphoryl]-L-alanyl-L-proline (5), and N alpha-[(2-phenylethyl)phenoxyphosphoryl]-L-alanyl-L-proline (12) was determined spectrophotometrically at pH 7.5 and 37 degrees C in both Tris and phosphate buffers. These moderately potent inhibitors of angiotensin converting enzyme (Ki greater than 0.8 microM) all hydrolyze, losing 1 mol of phenol to yield highly potent inhibitors (Ki = 0.5-18 nM). The half-times for loss of 1 mol of phenol in Tris buffer are 22 days (2), 3.4 h (5), and 21 days (12). The half-times in phosphate buffer were not significantly different. The mono(4-nitrophenoxy) ester 6 (Ki = 18 nM) loses its 1 mol of nitrophenol with a half-time of 35 h to yield N alpha-phosphoryl-L-alanyl-L-proline 16 (Ki = 1.4 nM), which hydrolyzes at the P-N bond with a half-time of 2.2 h. Hydrolysis of the P-N bond in 2 and 12 was not observed during the time course of the kinetic experiments. The two phosphoramidate diesters 2 and 5 and the phosphonamidate monoester 12 thus release powerful inhibitors of angiotensin converting enzyme with a known time course at physiological pH and temperature in vitro. A time-dependent increase in inhibitory potency against converting enzyme that paralleled the kinetics of phenyl ester hydrolysis was confirmed in vitro.