The discovery of spherical accumulations of iron oxide mineralization on Mars has prompted renewed interest in widespread but enigmatic accumulations of iron oxide on Earth. The rinded iron oxide concretions found in the Navajo Sandstone (Utah, USA) have been among the most extensively studied of these occurrences. At least four different models have been proposed for the formation of iron oxide concretions in the Navajo Sandstone. These include 1) mixing of reduced Fe(II)-bearing and oxygenated groundwaters, 2) neutralization of oxidized, acidic waters by calcite concretions, 3) neutralization of reduced, acidic waters by calcite concretions, and 4) oxidation of siderite by microbes during invasion of the Navajo aquifer by oxygenated groundwaters. We measured δ56Fe values of iron-bearing minerals from whole-rock Navajo Sandstone, concretionary iron oxide cements, iron oxide stains that are oriented in the direction of paleo-groundwater flow, iron oxide stains on the interior of rinded concretions, and ferroan carbonate cements. We have then combined these data with published measurements to evaluate each of the four published models. Whole rock Navajo Sandstone from the Kolob Plateau has been interpreted to be unaltered by bleaching fluids and has a δ56Fe value of 0.21‰. Iron in the Kolob Plateau sample comprises a mixture of hematite grain coatings (δ56Fe = 0.12‰) and a magnetic mineral fraction (interpreted to comprise largely detrital magnetite) that has a higher δ56Fe value (0.45‰). Iron oxide cements have δ56Fe values that are typically negative and as low as −0.74‰. In contrast, iron oxide-stained Navajo Sandstone in close proximity to a rinded iron oxide concretion has δ56Fe values that are 0.9 to 1‰ greater than the δ56Fe value of the associated iron oxide concretion. Finally, ferroan carbonates from the Navajo Sandstone yield δ56Fe values as low as −0.86‰. These values are in good agreement with measurements published previously for lithologies for which comparable measurements have been made. The presence of ferroan carbonates with negative δ56Fe values is evidence that aqueous Fe(II) was transported through the Navajo Sandstone aquifer and produced carbonate mineralization with negative δ56Fe values. The difference in δ56Fe values between the iron oxide concretions and associated iron oxide staining is more consistent with equilibrium fractionation between aqueous Fe(II) and Fe(II) adsorbed on quartz grains than it is with equilibrium fractionation between aqueous Fe(II) and any reduced or oxidized iron oxide mineral species. The negative δ56Fe values of the iron oxide concretions are best explained by oxidation of a reduced iron phase in a closed system from which no aqueous Fe(II) could escape. We interpret the iron isotope data as evidence that iron was stripped from Navajo sand grains by reducing fluids, reprecipitated in the Navajo aquifer as siderite, and oxidized by groundwaters under closed conditions during incision of the Colorado Plateau.