This work was carried out to determine the effect of 2,4-dinitrophenol(DNP) on in vivo nitrate reductase activity in the roots of 6 day old rye (Secale cereale L.) seedlings. The nitrate reductase activity in the roots of 6 day old rye seedlings pretreated with 0.5 mM DNP was higher than that of the control group in all the experimental conditions. The optimal concentration of KNO_3 for maximum nitrate reductase activity was 10 mM in both control and treated group. The nitrate reductase activity in the treatment of 10 mM KNO_3 gradually increased for 4 h in both groups, and then maintained constantly. The nitrate reductase activity occurred per hour was highest at 1 h in both groups, while it was declined by large degrees as time goes on. The daily pattern of nitrate reductase activity was gradually decreased in both groups with the passage of day. The optimal pH for nitrate reductase activity was 7.5 in both groups. From the results of this experiment and a previous paper(Kwon et al., 1991), it was determined that the nitrate reductase activity in both roots and shoots of rye seedlings was increased by the treatment of 0.5 mM DNP, and particulary in both grous, the nitrate reductase activity in the roots of rye seedlings was higher than that in shoots of them.