Drought stress affects plant photosynthesis and transpiration, as well as soil respiration and evaporation. In a coupled plant and soil system, drought can strongly impact the exchange of 18O in CO2 between the ecosystem and atmosphere. In this study we present diurnally resolved measurements of d18O values in ecosystem water pools in a sorghum field in the ARMCART SGP region (Oklahoma, USA). Over a 4-day period we measured continuous ecosystem CO2 and H2O fluxes using eddy correlation; soil moisture and temperature; d18O of soil water in 4 soil layers, leaves, and stems 4 times per day; and 18O in H2O at 2 heights above the plant canopy. Ecosystem CO2 fluxes reflect the impact of midday water stress. Measured soil water d18O values showed strong diurnal patterns reflecting soil-surface evaporation during the day and recharge from deeper soil layers at night. Diurnal soil water d18O values in the top soil layers varied by up to 6 . The d18O values of stemwater also varied over the course of the day, but to a smaller extent. Leaf water d18O values increased by up to 10 over the day. To interpret these data and to estimate C18OO ecosystem fluxes we applied a mechanistic model, called ISOLSM, which simulates H218O and C18OO ecosystem stocks and fluxes between ecosystems and the atmosphere. ISOLSM includes modules to compute canopy vapor, leaf water, and vertically resolved soil water H218O content; leaf photosynthetic and retro-diffusive fluxes of C18OO; root and microbial production of CO2; soil diffusive fluxes of CO2 and C18OO and equilibration of CO2 with 18O in soil water; and abiotic soil exchanges of C18OO. The model has been tested in a C4 dominated tallgrass prairie site close to the field studied here. Drought stress strongly affected the variability of the 18O content of near-surface soil water. The low soil moisture levels impacted the soil-surface C18OO fluxes via interactions with the soil-gas diffusion coefficient, microbial and root CO2 production, and the heavy near-surface soil water. Drought stress also impacted stomatal conductance, which in turn affected transpiration, the canopy air space vapor and vapor 18O content, and leaf C18OO exchange. Finally, we present a sensitivity analysis of the ecosystem C18OO exchange to the method used to quantify the impacts of plant water stress.