We made use of land cover maps, and land use change associated with urbanization, to provide estimates of the loss of natural resource lands (forest, agriculture, and wetland areas) across the 168,000 km2 Chesapeake Bay watershed. We conducted extensive accuracy assessments of the satellite-derived maps, most of which were produced by us using widely available multitemporal Landsat imagery. The change in urbanization was derived from impervious surface area maps (the built environment) for 1990 and 2000, from which we estimated the loss of resource lands that occurred during this decade. Within the watershed, we observed a 61% increase in developed land (from 5,177 to 8,363 km2). Most of this new development (64%) occurred on agricultural and grasslands, whereas 33% occurred on forested land. Some smaller municipalities lost as much as 17% of their forest lands and 36% of their agricultural lands to development, although in the outlying counties losses ranged from 0% to 1.4% for forests and 0% to 2.6% for agriculture. Fast-growing urban areas surrounded by forested land experienced the most loss of forest to impervious surfaces. These estimates could be used for the monitoring of the impacts of development across the Chesapeake Bay watershed, and the approach has utility for other regions nationwide. In turn, the results and the approach can help jurisdictions set goals for resource land protection and acquisition that are consistent with regional restoration goals.