Estimation of hæmatin iron and the oxidation-reduction equivalent of cytochrome c

Abstract

In previous communications it has hen shown that tire component c of cytochrome belongs to tire hæmochromogen class of compounds (Keilin, 1925, 1930); topical hæmochromogen being composed of a bæm (Fe'-porphyrin) united to a nitrogenous substance. The oxidation-reduction equivalent of hydrazin protohæmochromogen was found by Haurowitz (1927) to be equal to one equivalent of hydrogen per iron atom of the pigment. In the present study the oxidation-reduction equivalent of cytochrome c , extracted from baker's yeast, has been determined by titration with a ferrous salt in presence of alkaline tartrate, a method which was shown to be suitable for the titration of hæmatin and methæmoglobin (Hill and Holden, 1927). For this purpose it was essential to determine the concentration of cytochrome in the preparation of this pigment expressed in terms of its hæmatin. This was achieved in two different ways: (1) by comparing the colour and the absorption spectrum of reduced cytochrome with those of pyridine hæmochromogen prepared from a known concentration of mesohæmin, the absorption spectrum and colour of which match well those of cytochrome; and (2) by the estimation of iron in organic combination, these two methods showed that the hæmatin iron was equal to the total iron, while the free or unmasked iron in our preparations of cytochrome was found to be quite negligible.