The Quantitative Determination of Cytochrome C*

In the course of a quantitative study of the distribution of cytochrome c in cancer tissue a consideration of the available methods of analysis led to the development of an improved method which has been applied to this particular study.l The method has been used under such a variety of conditions that it is believed that it will give satisfactory results if applied to other problems involving the concentration of cytochrome c in animal tissues. Both manometric and spectrophotometric techniques have been applied to the determination of cytochrome c by previous investigators. Von Euler and Hellstrom (2) reported the cytochrome c content of Jensen rat sarcoma on the basis of a qualitative spectrophotometric method. Junowicz-Kocholaty and Hogness (3) have presented a method for cytochrome c determination which is essentially Keilin and Hartree’s method for the preparation of pure cytochrome c (4) placed on a quantitative basis. The method requires the use of 100 gm. of tissue, a quantity too large for experiments in which rats and mice are used as experimental animals. Fujita, Hata, and Numata (5) presented a spectrophotometric method for the determination of cytochrome c in which an attempt was made to isolate the cytochrome in pure form. Attempts to recover added cytochrome c by this method gave unsatisfactory results. It is believed that an incomplete precipitation of cytochrome c occurred during the acetone treatment. By far the most satisfactory method in the literature thus far is