These data consist of stream water chemistry for selected Welsh upland rivers. The sampling sites were located in sixty one small and medium catchments. Catchments were chosen from the Welsh Acid Water Surveys (WAWS) program (41 sites) and the Wye catchment (20 sites). Results for pH, alkalinity, conductivity and major cation and anion measurements are presented for the WAWS catchments. Results for pH, alkalinity, conductivity and major anion measurements are presented for the Wye catchment. Samples from the Wye catchment were collected in May 2012. Samples from the WAWS catchments were taken during the summer and autumn of 2012 and spring and summer of 2013. The data were collected to characterise water chemistry variation along a gradient of aquatic biodiversity associated with different environmental settings for example land-use intensify and recovery from acidification. Dr Isabelle Durance was responsible for organising the surveys, Dr Hugh Feeley was in charge of collecting and preserving the water samples. Analysis of the water samples was carried out at the Forest Research Laboratories. The work was carried out under Diversity in Upland Rivers for Ecosystem Service Sustainability (DURESS) project (Grant reference NERC NE/J014818/1). DURESS was a project funded by the Natural Environment Research Council (NERC) Biodiversity and Ecosystem Service Sustainability (BESS) programme. Full details about this dataset can be found at https://doi.org/10.5285/d329ca18-c3d8-49f0-b2fd-5243d76dc650