Studies on the Chromatographic Separation of Hemin a and Protohemin from Heart Muscle
1965 年 58 巻 1 号 p. 39-46
詳細
1965 年 58 巻 1 号 p. 39-46
1. Column- and paper-chromatographic methods for the separation of hemin a and protohemin from heart muscle extracts were described. Silicic acid impregnated paper and alumina impregnated paper were used for paper chromatography. Alumina was used for column chromatography.
2. Hemin a and protohemin could be separated effectively on silicic acid impregna-ted paper with ethanol-water-formic acid or acetone-water-formic acid as the solvent system. Effective separation was also observed on alumina impregnated paper with ethanol-water-formic acid as the solvent system. In all cases, the Rf value of hemin a was higher than that of protohemin.
3. There was essentially no tailing with these methods. The degree of separation was dependent on the concentrations of both water and formic acid.
4. A separation was observed on alumina impregnated paper with chloroform-formic acid or chloroform-acetic acid as the solvent system. However, in these cases, the Rf values of hemin a were smaller than those of protohemin.
5. Hemin a was separated on a alumina column, using a acetone-water-formic acid mixture as eluant. Hemin a was eluted prior to protohemin by this method. However, as the hemin a fraction and the protohemin frac-tion overlapped considerably, rechromato-graphy was necessary to obtain pure hemin a.
6. The spectrophotometrical properties of hemin a obtained by this method were identical with those of hemin a from purified cyto-chrome c oxidase and also agreed with the descriptions of many previous investigators. But no substance corresponding to the hemin A reported by Caughey and York was found during this work.
The authors express their deep gratitude to Dr. S. Horie of this Department for his valuable advice.
