We present a novel approach to study the sustainability of ancient Mediterranean agriculture that combines the measurement of carbon isotope discrimination (Δ¹³C) and nitrogen isotope composition (δ¹⁵N) along with the assessment of quality traits in fossil cereal grains. Charred grains of naked wheat and barley were recovered in Los Castillejos, an archaeological site in SE Spain, with a continuous occupation of ca. 1500 years starting soon after the origin of agriculture (ca. 4000 BCE) in the region. Crop water status and yield were estimated from Δ¹³C and soil fertility and management practices were assessed from the δ¹⁵N and N content of grains. The original grain weight was inferred from grain dimensions and grain N content was assessed after correcting N concentration for the effect of carbonisation. Estimated water conditions (i.e. rainfall) during crop growth remained constant for the entire period. However, the grain size and grain yield decreased progressively during the first millennium after the onset of agriculture, regardless of the species, with only a slight recovery afterwards. Minimum δ¹⁵N values and grain N content were also recorded in the later periods of site occupation. Our results indicate a progressive loss of soil fertility, even when the amount of precipitation remained steady, thereby indicating the unsustainable nature of early agriculture at this site in the Western Mediterranean Basin. In addition, several findings suggest that barley and wheat were cultivated separately, the former being restricted to marginal areas, coinciding with an increased focus on wheat cultivation. Copyright © 2008 John Wiley & Sons, Ltd.