Scavenging of hydroxyl radicals but not of peroxynitrite by inhibitors and substrates of nitric oxide synthases

Abstract

1. The nitric oxide synthase inhibitor N^G-nitro-L-arginine methyl ester (L-NAME) is widely used to study the role of NO^• in physiological and pathological processes, including its role in the generation of the cytotoxic species peroxynitrite (ONOO⁻) and of reactive oxygen radicals such as hydroxyl (OH^•). Often L-NAME is applied to tissues at mM concentrations. At such high concentrations, it might act as a free radical scavenger. A similar possibility might apply to the use of high levels of arginine to study the role of NO^. in atherogenesis.

2. We therefore examined the rate of scavenging of OH^• by L-NAME and found that L-NAME reacts more quickly with OH^. than the established ‘OH^. scavenger' mannitol and the widely used `OH^• trap' salicylate. However, D-NAME can scavenge OH^• at rates equal to L-NAME. Both L- and D-arginine were also good OH^• scavengers, comparable in effectiveness to mannitol.

3. Neither L-NAME, D-NAME, L-arginine nor D-arginine was able to inhibit ONOO⁻-dependent nitration of tyrosine, suggesting that they are unlikely to be scavengers of ONOO⁻-derived nitrating species.

4. Neither L-NAME, D-NAME, L-arginine nor D-arginine was able to inhibit the inactivation of α₁-antiproteinase by ONOO⁻, suggesting that they cannot prevent direct oxidations by peroxynitrite.

5. We conclude that L-NAME has sufficient activity as an OH^• scavenger to confound certain pharmacological experiments. However, this explanation of its biological effects can be ruled out if control experiments show that D-NAME has no effect and that L-arginine (also a free radical scavenger) antagonizes the action of L-NAME.

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