Electrolyte pH tremendously affects the electricity output of microbial fuel cells. However, its underlying molecular mechanism remains elusive, in particular for Shewanella oneidensis MR-1, one of the most widely adopted electrogenic microorganisms. Herein, we found that MFCs were able to deliver a significant (but different) electricity output in a wide range of pH (from 6 to 10), with the maximum at pH = 9 (alkaline), which delivers similar to 1.5 times' higher power output than that at pH = 7 (neutral). Furthermore, cyclic voltammetry analysis showed an enhanced electrochemical activity of riboflavin (responsible for extracellular electron transfer of Shewanella) at alkaline pH. Strikingly, the concentration of riboflavin synthesized by Shewanella in MFCs at different pH showed a good correlation with the electricity output of MFCs. Thus, our results substantiated that the increase of riboflavin biosynthesis by Shewanella at the alkaline condition underlies the improvement of the electricity output in MFCs. (C) 2012 Elsevier Ltd. All rights reserved.