Disruption of natural flow regimes, nutrient pollution, and other consequences of human population growth and development have impacted most major rivers of the world. Alarming losses of aquatic biodiversity and biotic homogenization coincide with human-caused river alteration, but effects on aquatic ecosystem processes are not as well documented. This is because unaltered systems for comparison are scarce, and some ecosystem-wide effects may take decades to manifest. We evaluated aquatic ecosystem responses to extensive river-floodplain engineering and nutrient addition in the Rio Grande of southwestern North America as revealed by changes in trophic structure of, and resource availability to, the fish community. Stable Isotope Analysis (SIA) was conducted on museum-preserved fishes collected over a 70-year period of intensive river management and exponential human population growth. Trophic complexity and resource heterogeneity for fish consumers (measured as 'isotopic niche breadth') decreased following sediment deprivation and channelization, and these effects persist into the present. Increased nutrient inputs led to δ15N enrichment in the entire fish community at all affected sites, and a shift to autochthonous sources of carbon at the most proximal site downstream of wastewater release probably via 'bottom-up' transfer. Overall, retrospective SIA of apex consumers suggests radical change and functional impairment of a floodplain river ecosystem already marked by significant biodiversity loss.