The relation of diaphorase of human erythrocytes to inheritance of methemoglobinemia.

Hereditary methemoglobinemia has been found to have a relatively high prevalence in Alaskan Eskimos and Indians (1). Persons with this condition have been found to have a complete lack of diphosphopyridine nucleotide diaphorase activity in red cells (2). Of the 21 known cases of methemoglobinemia in 13 families in Alaska, 12 cases in 7 families occurred in Ingalik Indians, a small group of about 800 Athabascans living on the Innoko, Yukon and Kuskokwim Rivers. The present study is concerned with a study of diaphorase levels of red cells of 271 of these people and of 47 Eskimos living nearby. Evidence is presented which indicates that parents of methemoglobinemic children have about half the normal level of erythrocyte diaphorase, and that the condition is inherited as a simple recessive trait. Mlalloy (3) that an optical density of 0.2 at 600 mAu in a 10 mm cell was given by 1.08 mg methemoglobin per ml. To each of two cuvets were added 0.05 ml 1 NI Tris (hydroxymethyl)aminomethane (pH 7.55), 0.1 ml 0.01 M sodium ethylenediamine tetraacetate (EDTA) and 0.05 ml 0.0012 M 2,6-dichlorobenzenoneindophenol (Eastman). To one cuvet was added hemolysate equivalent to 3.25 mg metheinoglobin and water to a volume of 2.98 ml; to the blanik was added 2.78 ml water. The reaction was started by addition of 0.02 ml 0.0088 M reduced diphosphopyridine nucleotide (DPNH). Optical density in a 10 mm cell was measured at 3-minute intervals for 21 minutes at 600 my in a Beckman spectrophotometer. The spectrophotometer was equipped with a water jacket, through which water at 250 C was circulated. Under these conditions, the decrease in optical density both with hemolysate and. in the blank was nearly linear. The level of diaphorase activity of the hemolysate wvas taken to be the rate of reaction expressed as change in absorbance per miniute (5A per miniute) corrected for the blank.