Responses of near-surface permafrost and glacial ice to climate change are of particular significance for understanding long-term effects on global carbon cycling and carbon export by high-latitude northern rivers. Here we report Δ14C-dissolved organic carbon (DOC) values and dissolved organic matter optical data for the Yukon River, 15 tributaries of the Yukon River, glacial meltwater, and groundwater and soil water end-member sources draining to the Yukon River, with the goal of assessing mobilization of aged DOC within the watershed. Ancient DOC was associated with glacial meltwater and groundwater sources. In contrast, DOC from watersheds dominated by peat soils and underlain by permafrost was typically enriched in Δ14C indicating that degradation of ancient carbon stores is currently not occurring at large enough scales to quantitatively influence bulk DOC exports from those landscapes. On an annual basis, DOC exported was predominantly modern during the spring period throughout the Yukon River basin and became older through summer-fall and winter periods, suggesting that contributions of older DOC from soils, glacial meltwaters, and groundwater are significant during these months. Our data indicate that rapidly receding glaciers and increasing groundwater inputs will likely result in greater contributions of older DOC in the Yukon River and its tributaries in coming decades.