Six genotypes of winter wheat (Triticwm aestivum L.) differing in grain protein concentration were grown on a nutrient solution containing low concentrations of N03-(2 millilar). Total N03-uptake varied between genotypes but was not related to grain protein content. An in vivo nitrate reductase assay was used to determine the affinity of the enzyme for NOs-, and large phenotypic variations were observed. In vivo estimations of the concentration and size of the metabolic pool were variable. However, the three genotypes with the higher ratios of metabolic pool size to leaf total N03-concentration were the high protein varieties. It is proposed that a high affinity of nitrate reductase for nitrate might be a biochemical marker for the capacity of the plant to continue assimilating N03-for a longer period during the last stage of growth.The potential use of such physiological criteria as markers is discussed, in particular with respect to breeding programs for the development of plants with efficient nitrogen utilization.To improve the efficiency of nitrogen fertilization and reduce its associated problems (eutrophication, increasing costs), more information is required on the processes of nitrogen utilization by plants.Use of biochemical and/or physiological criteria as an aid in breeding programs for cereal improvement has been proposed (8). Among the biochemical markers for improved efficiency of N utilization are NO3-uptake (7, 18), NRA3, total N content, and percentage in straw (13), harvest N index (15), and leaf protease activity (16). NRA has been widely studied (8,18). Nevertheless, results were not always sufficiently conclusive to justify the proposal of NRA as a good criterion for predicting a high efficiency of N utilization (e.g. high grain protein concentration) (8).Of the methods used for the evaluation of NRA in plants, the in vivo assay has often been preferred to the in vitro assay because of its simplicity. It has been shown that in vivo NRA determinations without exogenous N03 added (NRAa), give a good approximation of the actual amount of N reduced (21). It has been proposed that leaf N03-is distributed in two compartments: one small, rapidly exchangeable, cytoplasmic metabolic pool; and a large, vacuolar storage pool (9). The N03 in the metabolic pool is thought to control NR induction and NRAa (1, 9). Ferrari et al. (9) have proposed a method for the estimation of the MPS, but no attempts have been made either to determine the variations of MPS between genotypes or to estimate the MPC of N03.Skiba et aL (22) have estimated the apparent Km of NR for N03 in wheat and concluded that, with their in vivo determination, there was a good correlation between the grain protein concentration and the affinity of NR for NO3 among different genotypes, especially at low N fertilization levels. However, it was shown later that K+ ions had a stimulatory effect on N02 accumulation during the in vivo NR assay (11). Therefore, K+ concentration must be kept constant when varying the N03 concentration in ...