DETERMINATION OF SERUM IRON AND PSEUDOHEMO- GLOBIN IRON WITH o-PHENANTHROLINE

In a recent publication (1) we reported results obtained by the use of a new analytical procedure for iron in blood plasma. This method, which we find to be more sensitive than those which we have described previously (2-4), depends upon the photoelectric measurement of the red color of the complex formed by ferrous iron with o-phenanthroline. This reaction was first applied, as far as we know, to the determination of small amounts of iron by Saywell and Cunningham (5). Different aspects of the reaction have been studied by Hummel and Willard (6), Fortune and Mellon (7), Thiel, Heinrich, and van Hengel (8), and others. Heilmeyer and Plotner (9) were the first to apply o-phenanthroline to the measurement of iron in blood serum. Reagents~ 1. Hydrochloric acid, 1.2 per cent solution in water. 2. Hydrochloric acid, 0.3 per cent solution in water. 3. Trichloroacetic acid, 20 per cent solution in water. 4. Sodium acetate, saturated solution in water. 5. 2 M acetate buffer, pH 4.5, prepared by mixing 90 volumes of 2 M sodium acetate and 110 volumes of 2 M acetic acid. 6. Hydrazine sulfate, 1 per cent solution in 2 M acetate buffer, pH 4.5; this solution must be freshly prepared daily. 7. o-Phenanthroline monohydrate, 0.1 per cent solution in water; the preparation may be warmed to accelerate solution. It should be rejected if any color appears. 8. A blank solution prepared by mixing 1 volume of Reagent 1, 1 volume of Reagent 3, and 2 volumes of water. The blank