ABSTRACT Different depositional environments (fluvial, deltaic, lacustrine, tidal-marine) have been proposed for the Upper Miocene to Pliocene Madre de Dios Formation exposed in the upper reaches of the Amazon River catchment in the Andean retroforeland region. This study constrains the stratigraphy, depositional environment, and drainage evolution in southwestern Amazonia through petrographic and provenance analysis of the sand and mud fractions of the three recognized members (A–C) of the Madre de Dios Formation at three stratigraphic sections measured previously along riverbank outcrops: Cerro Colorado, Piedras River, and Candelaria. Petrographic analyses of thin sections of sand separates from 32 sandy samples showed them to be litho-quartzose to quartzo-lithic in composition, with variable feldspar content and a recycled-orogen provenance. Sand components were predominantly monomineralic to polymineralic quartz, and sedimentary and metamorphic lithic fragments. Muscovite, potassium feldspar, plagioclase, and volcanic lithics were less abundant. These sand components are consistent with derivation from the Andean range to the west. Quartz and feldspar content generally increased up section from member A to member B. Sand composition in member C is similar to the modern river sand composition, consistent with recycling of Madre de Dios Formation sand into the modern river system. Petrographic analyses of 144 smear slides of the mud fractions showed no significant changes in silt composition, i.e., mainly quartz, feldspar, and mica, among the three members. X-ray diffraction of eight mud samples showed their clay mineralogy to be dominated by kaolinite and illite, with some smectite and chlorite in member B. None of the 144 smear slides or the 32 thin sections contained marine or marginal-marine biogenic debris. Mud-rich samples from the Madre de Dios Formation exhibit six main colors that characterize distinct intervals (designated I to VI) that occur in the same stratigraphic order in each measured section, from I at the base to VI at the top. The boundaries of these intervals do not correspond directly to member (A, B, C) boundaries; therefore, the colors are at least partly secondary. If primary, the red to orange to brown mudstone (which is dominant in Madre de Dios Formation members A and C) would suggest development in oxidizing environments, consistent with fluvial systems. Based on its light olive-gray color and smectite content, interval IV in member B may have been deposited in, or subjected to, a more reducing environment, such as a lacustrine-deltaic setting, with low-lying topography and poor drainage. In sum, the Madre de Dios Formation exhibits up-section compositional and thickness trends that are consistent with changes in depositional environment from fluvial (member A) to lacustrine/deltaic (member B) to fluvial (member C), as proposed by previous workers.