In this study there was evaluated the influence of the initial relation of Fe3+/Fe2+ in a process of biodesulfurization of a coal sample. We used a culture medium and a strain of Acidithiobacillus ferrooxidans , with an initial concentration of 1200 mg/L total iron, varying the Fe3+/Fe2+ ratio between 80:20, 50:50 and 20:80. It was used a particle size minus 60 mesh (dp<246 μm). The process was monitored by periodic measures of pH, redox potential, concentration of total iron, Fe3+ and Fe2+ in solution. According to the obtained results, experiments using proportions Fe3+/Fe2+ of 50:50 and 20:80 showed better efficiency, presenting the highest percentages of bioleaching of iron in solution, reaching respectively 65% and 74% of oxidation of pyrite, after 14 days of process.