Bacterial electroactive biofilms attract researchers’ attention due to their ability for generation of electric current when electrons travel for very long, up to several tens microns, distance. It was demonstrated that this electron transfer (ET) ability correlates with the accumulation in the BF huge amounts of a c-type cytochrome (Cyt), but molecular nature of this Cyt, its spatial localization, interaction with the environment, regulation and the controlling factors are not completely understood. Bacterial cell polarity is an internal cell asymmetry that correlates with the cell ability to sense energy and metabolite sources, chemical signals, quorum signals, toxins, and movement in the desired directions. This ability also plays central role in cell attachment to various surfaces and biofilm formation. Biochemical analysis and molecular imaging demonstrate that sensing of energy and nutrients by the cell correlates with asymmetric protein, lipid and other molecule distributions within the cell. In the present work, we develop a new experimental approach for analyzing redox potential spatial distribution within individual bacterial cells and use this approach for identification how attachment to inorganic mineral, mica affects redox state inside individual G. sulfurreducens cells. The approach is based on confocal Raman microscopy allowing for precise estimation of redox state and localization of subcellular cytochromes. Applying Gaussian deconvolution and principal component analysis we are able to identify spectral signatures of cytochromes in different redox states and intracellular locations. Our results give a new tool for quick identification of internal and surface exposed cytochromes and their localization at bacterial cell sides facing the substrate and the solution. They open up the possibility for analyzing electrochemical potentials inside individual bacterial cells and indicate that Cyt redox state might be a driving force switching cell metabolic activity that leads to irreversible cell attachment to electrode, cell-to-cell association, and biofilm formation.