Abstract.The significance of blue light‐stimulated stomatal conductance for carbon assimilation (A), stomatal conductance (g), intercellular CO2(Ci), stomatal limitation of A (L), transpiration (E) and water use efficiency (W = A/E), was determined in a C4and a C3species. W and L were evaluated for steady‐state gas exchange with constant, saturating red light (As, gs, Es), and for the integrated gas exchange above the steady state baseline induced by a single, brief pulse of blue light (Ap, gp, Ep). Sugarcane (Saccharumspp. hybrid), a C4grass, and soybean (Glycine max) a C3dicot, were compared. Sugarcane exhibited typical C4behaviour, with A saturing at Ciof ca. 200 μmol mol−1, compared to>500 μmol mol−1in soybean. Steady‐state W was also considerably higher in sugarcane. The extent of stomatal opening in response to a blue light pulse, from baseline (gs) to the maximum value of conductance during the opening response (gm), was similar in the two species. More rapid opening and closing of stomata in sugarcane resulted in a smaller integrated magnitude of the conductance response (gp) than in soybean. At the peak of the blue light response, both species exhibited similar levels of L. During the response to the pulse of blue light, A and Ciincreased and L decreased to a greater extent in sugarcane than in soybean. As a result, the gas exchange attributed to the stomatal response to blue light exhibited a higher ratio of Apto Ep(Wp) in sugarcane than in soybean. This Wpwas lower in both species than was the Wsassociated with the steady state gas exchange. The two species did not differ in the rate of induction of photosynthetic utilization of elevated Ci. The greater stimulation of A in sugarcane was attributed to its C4attributes of greater carboxylation efficiency (slope of the A versus Cirelationship), lower gsand prevailing Ci,s, and greater Lsunder steady‐state red illumination. Despite saturation of A at low levels of Ciin C4species, the gas exchange attributed to the stomatal response to blue light decreased L and contributed considerably to carbon acquisition, while maintaining the high level of W associated with