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Abstract Nitrate nitrogen (NO3 −_N) leaching increased with nitrogen (N) fertilization under high water supply to the field negatively affected the maize growth and performance. This study aimed to understand the mechanisms of NO3 −_N leaching on a biochemical basis and its relationship with plant performance with 5 different doses (0, 200, 250, 300, 350 kg N ha− 1) of N fertilizers under low (60%; LW) and high (80%; HW) water holding capacity. Soil and plant enzymes were observed at different growth stages (V9, R1, R3, and R6) of the maize, whereas the leachates were collected at 10-days intervals from the sowing date. The LW had 10.15% lower NO3 −_N leachate than HW, with correspondence increases in grain yield (25.57%), shoot (17.57%) and root (28.67%) dry matter. Irrespective of the irrigation water, RubisCo, glutamine synthase (GS), nitrate reductase (NR), nitrite reductase (NiR), and glutamate synthase (GOGAT) activities increased with increasing N fertilizer up to the V9 growth stage and decreased with approaching the maturity stage (R6) in maize. In HW irrigation, soil total N, GOGAT, soil nitrate (NO3 −_N), leached nitrate (LNO3 −_N), root N (RN), leaf N (LN) were positively correlated with N factors suggesting the higher losses of N through leaching (11.3%) compared to LW irrigation. However, the malondialdehyde (MDA), hydrogen peroxide (H2O2), superoxide (O2 −), and proline were negatively correlated with the other enzymatic activities both under LW and HW irrigation. Thus, minimizing the NO3 −_N leaching is possibly correlated with the LW and N300 combination without compromising the yield benefit and improving enzyme activities.