Microbial Fuel Cells (MFCs) is an innovative technology able to both clean organic waste of domestic or industrial origin and produce renewable energy converting directly chemical energy stored in chemical bonds to electrical energy. Our findings showed that the rate of substrate consumption (such as glucose) by Saccharomyces Cerevisiae (a common baker’s yeast) in the anaerobic compartment of a dual chambered MFC presented a great potential to generate electrons in a microbial fuel cell. Methylene blue was used as the electronophore in the anode compartment, while oxygen and hydrogen peroxide were tested as electron acceptors in the cathode compartment. The experiment was performed in closed circuit configurations under different loads ranging for evaluating the performance of the MFC with different concentration of methylene blue and electron acceptor. The fuel cell mediated by methylene blue carried out a maximum power generation of 110 mWm-2 and generated a maximum open circuit voltage of 479 mV with a short circuit current of 1.17 Am-2 with hydrogen peroxide as electron acceptor. The different contribute on the cell output of the mediator and electron acceptor concentration in anode and cathode compartment have been studied.