Table 3

Kinetic constants for mechanism-based inactivation of the iNOS and nNOS isoforms by diverse 1-aminoguanidines and 1-amino-S-methylisothiourea

Compound	iNOS		nNOS
Compound	K_I	k_inact max	K_I /k_inact max	K_I	k_inact max	K_I /k_inact max
	mM	min⁻¹	mM/min	mM	min⁻¹	mM/min
Aminoguanidines
Unsubstituted	0.7	1.50	2.14	3.3	0.35	0.11
1-Methyl-	2.0	0.85	0.42	17.0	0.69	0.04
2-Methyl-	3.5	2.48	0.71	14.0	2.57	0.18
2-Ethyl	0.12	0.48	4.0	1.0	0.46	0.46
2-Hydroxy-	0.30	0.17	0.57	0.68	0.27	0.40
2-Phenyl	1.10	0.39	0.36	4.0	0.56	0.14
AMITU	1.10	0.75	0.68	0.3	0.73	2.43

Standard incubations to assess NO formation rate (as measured by the spectrophotometric determination of methemoglobin formation from oxyhemoglobin) were constructed as described in Experimental Procedures and the legend to figure 3. Incubations were conducted containing 100 μM arginine without or with the indicated agent at concentrations ranging from 0.03 to 30 mM. The time- and concentration-dependent decreases of activity (methemoglobin formation rate) were recorded. The data obtained were plotted in Kitz-Wilson format as the half-time of inactivation at each specified concentration of agent vs.the reciprocal of agent concentration. Linear plots were obtained for all agents, and the values of K_I andk_inact max were determined from the ordinal and abscissal intercepts as exemplified in figure 3.