Natural background level (NBL) is a prerequisite for identifying groundwater pollution and for the evaluation of pollution control effectiveness. However, due to regional differences of natural geogenic, hydrogeological, biological and atmospheric processes, together with long-term intense anthropogenic activities, it is no longer reasonable to assess NBL as a single value without including human impact. This paper shows the importance of regional division based on the knowledge of geology, hydrogeology and anthropogenic activities on the assessment of NBLs. In the study, the Guanzhong Basin, situated in central China with an area of 20,035 km2 was divided into ten groundwater environmental units (GEUs) based on hydrogeological structures and groundwater flow systems. A total of 509 shallow groundwater samples were collected throughout the basin. Based on samples from each GEU, NBLs of Na+, Ca2+, Mg2+, HCO3−, SO42−, Cl−, F−, Cr6+, TH and TDS were evaluated by using iterative 2-δ technique (2-δ), calculated distribution function (CDF) and Grubbs’ test. The results indicate the distinct characteristics of NBLs for the various GEUs: NBLs of Na+, SO42−, Cl−, HCO3−, F− and TDS north of the Wei River obviously exceeded those in the south; NBLs of most ions for GEU-V and GEU-X were highest north and south of the Wei River, respectively. Through detailed analyses of the natural and anthropogenic influences on NBLs, we found that the outcrops of Tertiary rocks in the Li mountains determine the highest NBLs of chemical components for GEU-X south of the Wei River; long-term irrigation is responsible for the highest NBLs of TDS, Na+, Mg2+, HCO3−, SO42− and Cl− for GEU-V; rapid urbanization in Xi’an (GEU-VIII) slows down the accumulation of chemical components (e.g. TDS) in shallow groundwater by reducing evaporation and mixing of shallow water with deep water through ground fissures. Assessment of NBLs on a regional scale lays the basis for identification of groundwater pollution and is helpful for the reasonable management and effective protection of groundwater resources.