A Rapid and Simple Colorimetry of Iron in Hemoglobin With Sodium Hypochlorite and Bathophenanthroline

Abstract

A rapid and sensitive technique for colorimetric determination of iron in hemoglobin which requires no deproteinization is described.
Iron (about 17 nmoles) in hemoglobin of 200μl of 100 times diluted blood is oxidized to F3+and released by adding 200μl of sodium hypochlorite (active chlorine: 0.28 mol/l).This released Fe³⁺is reduced to Fe²⁺by adding 300μl of 0.17 mol/l ascorbic acid.ln order to completely release and reduce iron, after standing for 5 minutes at a room temperature, the released Fe²⁺is determined by colorimetry with 300μl of 1.05 mmol/l bathophenanthroline at 535 nm.Simultaneousy, denatured protein should be dissolved by adding 500μl of 0.2mol/l tris-maleate buffer (pH 7.0).Thus this method (C.V.=2.15%) requires no deproteinization procedure.This method correlates enough with mineralization-bathophenanthroline method (y=1.005x-0.01, r=0.995, n=30).