The correlation between CO(2) assimilation and nitrate reduction in detached spinach (Spinacia oleracea L.) leaves was examined by measuring light-dependent changes in leaf nitrate levels in response to mild water stress and to artificially imposed CO(2) deficiency. The level of extractable nitrate reductase (NR) activity was also measured. The results are: (a) In the light, detached turgid spinach leaves reduced nitrate stored in the vacuoles of mesophyll cells at rates between 3 and 10 micromoles per milligram of chlorophyll per hour. Nitrate fed through the petiole was reduced at similar rates as storage nitrate. Nitrate reduction was accompanied by malate accumulation. (b) Under mild water stress which caused stomatal closure, nitrate reduction was prevented. The inhibition of nitrate reduction observed in water stressed leaves was reversed by external CO(2) concentrations (10-15%) high enough to overcome stomatal resistance. (c) Nitrate reduction was also inhibited when turgid leaves were kept in CO(2)-free air or at the CO(2)-compensation point or in nitrogen. (d) When leaves were illuminated in CO(2)-free air, activity of NR decreased rapidly. It increased again, when CO(2) was added back to the system. The half-time for a 50% change in activity was about 30 min. It thus appears that there is a rapid inactivation/activation mechanism of NR in leaves which couples nitrate reductase to net photosynthesis.