Simultaneous Quantitation of Zinc Protoporphyrin and Free Protoporphyrin in Erythrocytes by Acetone Extraction

Erythrocyte protoporphyrin content is increased in several human and veterinary disorders in which heme synthesis is disrupted. The nature of the defect determines whether zinc protoporphyrin, unchelated (free) protoporphyrin, or both are present in the erythrocytes. Extraction of the protoporphyrin into acetone/water (80/20 by vol) is a simple, rapid technique by which each protoporphyrin species is delivered into solution without removing zinc from zinc protoporphyrin. The ratio of zinc protoporphyrin to total protoporphyrin concentration correlated significantly with the ratio of fluorescence emission intensities at the zinc protoporphyrin peak and at an isosbestic point. This concentration ratio may be used with a quantitative measure of total erythrocyte protoporphyrin to define the absolute zinc protoporphyrin and free protoporphyrin content of erythrocytes.