Oxidative Conversion of Thiocyanate to Cyanide by Oxyhemoglobin during Acid Denaturation

Human oxyhemoglobin (Hb-O2) converts thiocyanate to cyanide under acidic conditions, in a hemoglobin and thiocyanate concentration-dependent manner. Other naturally occurring hemoglobin derivatives and hematin did not result in cyanide-producing activity. Pre-denaturation completely destroyed cyanide production. In the presence of 10% phosphoric acid, 12 μM cyanide was produced by 1 mM Hb-O2 (per heme) and 1 mM thiocyanate. This conversion was observed over a wide range of acidic pH levels at 50°C, with an optimal pH of 4. The production of cyanide under strong acidic conditions was completed within 1 s, while under weak acidic conditions (pH ∼ 5) the production gradually increased for half an hour. Under both conditions, the time course for cyanide production coincided with the disappearance of this converting activity. Thiocyanate conversion by acidified Hb-O2 was completely suppressed by the addition of ascorbic acid, significantly suppressed by superoxide anion radical (O2−) scavengers or thiourea, but not effected by scavengers of hydroxyl radical, singlet oxygen, and protein radical. In addition to acidification, other processes for denaturation of Hb-O2 resulted in detectable production of cyanide, e.g., heating, extraction by organic solvents, and incubation with denaturating or oxidative reagents. These data suggest that O2− generated during Hb-O2 denaturation and also a hydroperoxyl radical converted under acidic conditions are the oxidizing species that convert thiocyanate to cyanide.