Effect of cyanide on the reaction of nitroprusside with hemoglobin: relevance to cyanide interference with the biological activity of nitroprusside

Abstract

The reaction of sodium nitroprusside (SNP) with deoxyhemoglobin (Hb) results in two distinct EPR-detectable species, the one-electron-reduced nitroprusside ion [(CN)5FeNO]3- and nitrosylhemoglobin (HbNO). In the presence of excess cyanide (CN-) only the signal for [(CN)5FeNO]3- is observed. Thus, while free CN- does not interfere with Hb reduction of SNP, it prevents transfer of the NO moiety to Hb. Electrolytic reduction of SNP under similar conditions, however, leads to [(CN)5FeNO]3- and a small amount of [(CN)4FeNO]2- resulting from loss of the CN- trans to the NO. Excess free CN- shifts the equilibrium between these two species toward [(CN)5FeNO]3-, thereby reducing the concentration of [(CN)4FeNO]2-. Thus, [(CN)4FeNO]2- appears to be responsible for the transfer of NO to Hb. Consistent with this mechanism, both [(CN)5FeNO]3- and [(CN)4FeNO]2- are observed when SNP is added to erythrocyte lysates. Under these conditions HbNO is formed more rapidly due to the higher concentration of the latter species with the labile NO. This observation suggests that red blood cell constituents capable of binding CN- shift the equilibrium between the reduced SNP ions toward [(CN)4FeNO]2-. In the reaction of reduced glutathione (GSH) with SNP, [(CN)5FeNO]3- is formed as well as low concentrations of an EPR-detectable GSH-SNP adduct. Excess free CN- introduces a lag in the appearance of these signals, suggesting that GSH mediates SNP reduction by a different mechanism from that of Hb, although it too is inhibited by CN-. The CN- stabilization of [(CN)5FeNO]3-, the reduced SNP species lacking a labile NO moiety, probably accounts for the ability of CN- to block or reverse the biological effects of SNP on aortic strips and human blood platelets. This chemical interaction appears to meet many of the criteria for competitive antagonism. By comparison, the vasodilator 3-morpholinosydnoneimine, which is a metabolite of molsidomine and which has an alkyl cyano and an alkyl nitroxide group, releases NO by an entirely different mechanism since free CN- has no effect on its biological activity.