Determination of the Molar Absorption Coefficients of c-Type Cytochromes Using Optically Transparent Thin-Layer Electrodes

The simultaneous monitoring of both the absorbance and charge consumed in electrolysis in a precalibrated Optically Transparent Thin-Layer Electrode (OTTLE) cell was demonstrated as a precise method for determining the molar absorption coefficients of electron-transfer proteins. In this method, the procedure for a direct determination of the concentration of the protein could be omitted. The values of the molar absorption coefficients of c-type cytochromes at the α-peaks of reduced forms were determined by this method: 28.4 mM−1 cm−1 (1 M = 1 mol dm−3) at 550 nm for horse heart cytochrome c, 119 mM−1 cm−1 at 552 nm for cytochrome c3 from Desulfovibrio vulgaris, Miyazaki F, and 116 mM−1 cm−1 at 552 nm for cytochrome c3 from Desulfovibrio vulgaris, Hildenborough.