Land use and land cover change (LUCC) is an acknowledged cause of the current biodiversity crisis, but the link between LUCC and biodiversity conservation remains largely unknown at the regional scale, especially due to the traditional lack of consistent biodiversity data. We provide a methodological approach for assessing this link through defining a set of major pressures on biodiversity from LUCC and evaluating their extent, distribution, and association with a set of physical factors. The study was performed in the Metropolitan Region of Barcelona (MRB, NE of Spain) between 1956 and 2000. We generated a LUCC map for the time period, which was reclassified into a set of pressures on biodiversity (forestation, deforestation, crop abandonment, and urbanization). We then explored the association of these pressures with a set of physical factors using redundancy analysis (RDA). Pressures encompassed 38.8% of the MRB area. Urbanization and forestation were the dominating pressures, followed by crop abandonment and deforestation. RDA showed a significant distribution gradient of these pressures in relation to the studied physical factors: while forestation and deforestation are concentrated in remote mountain areas, urbanization mainly occurs in lowlands and especially on the coast, and close to previous urban centers and roads. Unchanged areas are concentrated in rainy and relatively remote mountain areas. Results also showed a dramatic loss of open habitats and of the traditional land use gradient, both featuring Mediterranean landscapes and extremely important for their biodiversity conservation. Implications of these results for biodiversity management are finally discussed.