The nitrite methemoglobin complex—Its significance in methemoglobin analyses and its possible role in methemoglobinemia

Abstract

Evidence consistent with the formation of a reversible complex between excess free nitrite ion and the ferric heme groups of methemoglobin is presented. The existence of this complex in nitrited red cells or in lysates in the presence of appropriate concentrations of free nitrite can lead to gross underestimation of methemoglobin levels by a common spectrophotometric method. Complex formation is manifested by a decrease in the absorption of methemoglobin solutions in the range of 600–650 mμ. This source of error can be eliminated in red cell suspensions simply by washing. Similarly, lysate solutions may be dialyzed or diluted to effect dissociation of the complex. Attempts were made to estimate the dissociation constant of this complex at 25°, pH 7.4, and physiologic ionic strength. Although unknown side reactions between methemoglobin and nitrite interfere in the spectrophotometric determination of this constant, its value is probably not greater than 3 mM. This estimate indicates that the complex must exist transiently in vivo when mice receive nitrite doses adequate to convert more than one-third of the circulating blood pigment to methemoglobin. Thus, complex formation may contribute to the unusually sustained methemoglobinemia produced in mice by nitrite.