Biodiversity in urban areas is affected by a multitude of stressors. In addition to physico-chemical stress factors, the native regional species pool can be greatly reduced in highly urbanized landscapes due to area loss and fragmentation. In this study, we investigated how macrophyte composition and diversity in urban water systems are limited by the regional species pool and local environmental conditions. Canonical correspondence analysis of the macrophyte species composition revealed that urban and semi-natural water systems differed and differences could be related to local abiotic variables such as pH and iron concentrations. Macrophytes in the semi-natural area were typical for slightly acid and oligotrophic conditions. In urban water systems, exotic species characteristic of eutrophic conditions were present. In the semi-natural areas, the number of macrophyte species exceeded the number of species expected from species-area relationships of artificial water bodies in rural areas. In urban areas, the number of macrophyte species was similar to artificial water systems in rural areas. Macrophyte species present in the study areas also were generally found within 20-30 km distance to the study area. Macrophyte species composition in urban water systems and semi-natural water systems appeared to be influenced by the regional species pool within approximately 30 km of the locations. Nevertheless, site limitation ultimately determined the local macrophyte species composition and diversity in urban water systems and in semi-natural water systems