The purple photosynthetic bacterium Rhodospirillum molischianum has a cytochrome c subunit bound to the reaction center (RC). Redox titration of the cytochrome c showed the presence of a high-potential heme peaked at 558 nm in the α-absorption band region, with an E m (redox midpoint potential) value of 390 mV, and three low-potential hemes peaked at 552 nm, with E m values below 100 mV. The extent of flash-induced membrane potential, measured by the carotenoid band shift in chromatophores, varied depending on which heme was photo-oxidized at the ambient redox potential. If the extent of membrane potential is proportional to the distance of electron movement, the heme with highest potential, c-558, is assumed to be located in the most distant position from the special pair of bacteriochlorophyll in Rs. molischianum. Kinetic measurements of photo-oxidations of hemes, however, suggested that the highest potential heme is at the nearest position in Rs. molischianum, as in the case of Rhodopseudomonas viridis. We propose a model in which the magnitude of the membrane potential depends not only on the distance of electron movement across the membrane but also on the dielectric property in the cytochrome subunit which can be largely affected by the distance of the redox centers from the protein-aqueous interface. According to this model, the highest potential heme, c-558, is estimated to be proximal to the special pair and also to the aqueous surface, so that the largest electrostatic potential change occurs upon its photo-oxidation.