The effect of NO on phosphotyrosine protein phosphatases (PTPases) has been investigated in vivo. NO production is induced in interferon-gamma and lipopolysaccharide stimulated RAW-264.7 macrophages as indicated by the increase of NO2- in the medium. Our results demonstrate an inhibition of p-nitrophenylphosphatase activity as a consequence of macrophages activation. Under the described experimental conditions, most of the hydrolysis of p-nitrophenylphosphate can be ascribed to the action of cellular PTPases. The presence of NG-mono-methyl-L-arginine, a specific inhibitor of NO synthase decreases the inactivation rate of both membrane-bound and soluble PTPases. This evidence further confirms the ability of NO to inactivate PTPases and suggests a possible role of NO in the regulation of cellular processes involving this class of phosphatases.