Measurement of Glucose-6-phosphate Dehydrogenase and 6-Phosphogluconate Dehydrogenase Activites in Erythrocytes by Use of a Centrifugal Analyzer

摘要

We describe an accurate automated method for measuring activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49) in erythrocytes with a centrifugal analyzer. Blood is collected in microhematocrit tubes, centrifuged, and the erythrocytes are lysed with digitonin. Glucose-6-phosphate dehydrogenase activity is determined by mixing the hemolysate with glucose-6-phosphate, NADP+, and 6-phosphogluconate dehydrogenase #{ 233} (EC 1.1.1.44) in triethanolamine—EDTA buffer at pH 7.6, and measuring the rate of NADPH production for 3 min. Under these conditions 2 mol of NADPH are produced per mole of glucose-6-phosphate oxidized, ensuring the accuracy of the method and increasing its sensitivity. Activity is referred to hemoglobin, measured as cyanmethemoglobin. Activity of glucose-6-phosphate dehydrogenase added to hemolysates was well accounted for. Results obtained by our method and by the method of Bishop [ J. Lab. Clin. Med. 68, 149 (1966)] are virtually identical. Our method requires a small amount of blood and is accurate and rapid; thus it is well suited for use in surveying large populations for glucose-6-phosphate dehydrogenase deficiency. A simple modification of the method may be used to determine the activity of 6-phosphogluconate dehydrogenase.