The present study aims at investigating land use changes (as one of the effective human factors on water systems) as well as its relation with water quality at spatial scales of the entire basin, sub-basin and defined buffers (10 and 15 km) in Zayandeh-Rud Basin, Isfahan, Iran. By means of supervised classification method along with maximum likelihood algorithm, it classifies the land use map into five categories, including agriculture, bare lands, urban areas, vegetation, and water. The research collects data for 11 water quality parameters in seven sampling stations of Zayandeh-Rud Basin in 2002, 2009, and 2015 from Isfahan Water and Sewerage Organization. Correlation analysis is then conducted to investigate the effect of land use changes on water quality at different spatial scales. Land use analysis in the entire basin shows that despite an increase in urban and agricultural lands from 2002 to 2015, bare lands, vegetation, and water covers have had a decreasing trend. Moreover, various land uses at different scales show some correlation with water quality parameters. The strongest correlations in this study belong to sub-basin scale. Therefore, it is recommended to use this spatial scale to investigate the relation between land use and water quality parameters