Arctic peat soils contain vast reserves of organic C and are largely anaerobic. However, anaerobic respiration, particularly the role of Fe(III) and humic substances as electron acceptors, is not well understood in such ecosystems. We investigated these processes in a drained thaw lake basin on the Arctic coastal plain near Barrow, Alaska. We measured concentrations of soluble Fe and other potential electron acceptors, described the microbial community, and performed experiments in the laboratory and field to measure net rates of Fe(III) reduction and the relationship of this process to C cycling. In most areas within the basin, aerobic conditions existed only in the upper few centimeters of soil, though oxygen penetrated deeper in raised areas, such as rims of ice wedge polygons. Concentrations of nitrate and sulfate in soil pore water were low or negligible. Soil pore water contained surprisingly high concentrations of Fe(II) and Fe(III), in the range of hundreds of μM, suggesting the presence of organic chelators. The solid phase contained substantial amounts of iron minerals, with a progressively reduced oxidation state throughout the growing season. The most abundant 16S rRNA sequence in our gene survey was closely related to the Fe(III)-reducing bacterium, Rhodoferax ferrireducens, and other sequences closely related to Fe-transforming bacteria were found. Field and laboratory incubations with soluble Fe(III) and the quinonic compound, AQDS (a common humic analog), stimulated respiration and verified that Fe(III) reduction occurs in these soils. We conclude that reduction of Fe(III) and humic substances are major metabolic pathways in this ecosystem.