Two nearly adjacent subcatchments, located in the Adirondack Mountains of New York State, US, with similar atmospheric inputs of N (0.6 kmol ha−1 yr−1), but markedly different stream water solute concentrations, provided a unique opportunity to evaluate the mechanisms causing this variation. Subcatchment 14 (S14) had much greater stream water Ca2+ and NO3− concentrations (851 and 73 μmolc L−1, respectively) than Subcatchment 15 (S15) (427 and 26 μmolc L−1, respectively). To elucidate factors affecting the variability in stream water concentrations, soil and forest floor samples from each subcatchment were analyzed for total elemental cations and extractable N species. Mineral soil samples were also analyzed for exchangeable cations. Tree species composition was characterized in each subcatchment and potential differences in land use history and hydrology were also assessed. Compared with S15, soils in S14 had significantly higher total elemental Ca2+ in the forest floor (380 vs. 84 μmol g−1), Bs horizon (e.g. 1361 vs. 576 μmol g−1) and C horizon (1340 vs. 717 μmol g−1). Exchangeable Ca2+ was also significantly higher in the mineral soil (64 μmol g−1 in S14 vs. 8 μmol g−1 in S15). Extractable NO3− was higher in S14 compared with S15 in both the forest floor (0.1 vs. 0.01 μmol g−1) and Bs horizon (0.2 vs. 0.07 μmol g−1) while extractable NH4+ was higher in S14 vs. S15 in the forest floor (7 vs. 5 μmol g−1). The total basal area of ‘base-rich indicator’ tree species (e.g. sugar maple, American basswood, eastern hophornbeam) was significantly greater in S14 compared with S15, which had species characteristic of sites with lower base concentrations (e.g. American beech and eastern white pine). The disparity in stream water Ca2+ and NO3−, concentrations and fluxes between S14 and S15 were explained by differences in tree species composition and soil properties rather than differences in land use or hydrology. The marked difference in soil Ca2+ concentrations in S14 vs. S15 corresponded to the higher stream water Ca2+ and the larger contribution of base-rich tree species to the overstory biomass in S14. Soil under such species is associated with higher net mineralization and nitrification and likely contributed to the higher NO3− concentrations in the drainage waters of S14 vs. S15. Studies investigating differences in spatial and temporal patterns of the effects of chronic N deposition on surface water chemistry need to account for changes in tree species composition and how vegetation composition is influenced by soil properties, as well as climatic and biotic changes.