The hydrologic cycle plays an important role in carbon cycling, due to the coupling of vapor release and CO2 uptake during photosynthesis. This coupling, expressed as Water Use Efficiency (WUE) or Transpiration Ratio, can provide an inexpensive alternative for estimating the Net Primary Productivity (NPP) of terrestrial ecosystems. The D/H and 18O/16O trends of river water in the Mississippi basin are mostly indistinguishable from those of precipitation. This, combined with isotopic mass balance relationships, suggests that direct evaporation of surface water is small and evapotranspiration (ET) flux from the basin therefore consists mostly of interception and transpiration, with interception approximated from field studies. The calculated water flux associated with transpiration is 1500.8 km3 (77.3% of the evapotranspiration flux). Utilizing the average WUE of 864 mol H2O for each mole of CO2, the NPP of the Mississippi River basin amounts to 1.16 Pg C/yr, similar to the model estimates of the heterotrophic soil respiration flux of 1.12 Pg C/yr. This does not favor the postulated existence of a major sink for atmospheric CO2 in the temperate Northern Hemispheric ecosystems of the conterminous United States, but due to uncertainties in the input parameters we cannot discount the possibility that these ecosystems act as a modest sink.