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The uptake of urea fertilizer (NDFF), applied with 150 kg nitrogen (N) ha−1, topdressed in five splits of 30 kg N ha−1 (30 N) each at 7, 26, 45, 70 and 83 days after transplanting (DAT) of rice (Oryza sativa L.), was investigated in an improved (Khangma Maap, KM) and a traditional (Janam, JN) cultivar in Bhutan highlands, using enriched 15N stable isotope. The treatments were arranged in a split–split plot design, with N fertilizer levels as main plots, cultivars as subplots and topdressing treatments as sub-subplots, with all the sub-subplots receiving the same dose except different timing of one split of enriched 15N to determine partial N fertilize use efficiency at each split dose. Although cultivar differences were not recorded in soil N accumulation and in total dry matter N, KM produced 21% higher grain yields compared to JN due to higher grain harvest index and partial factor productivity of N. Irrespective of the cultivars, topdressing timing had significant effects on NDFF, with highest mean N recovery (REN) of 29% of applied 30 N at 45 DAT during active tillering stage, resulting in mean NDFF total (grain + straw) uptake of 8.71 kg N ha−1 compared to least effective topdressing timing at 7 DAT with mean REN of 12% and NDFF total of 3.51 kg N ha−1. In similarity to topdressing at 45 DAT, topdressing at 70 DAT (panicle initiation stage) was equally effective with mean REN of 27% across the cultivars. Hence, fertilizer N topdressing recommendations that combine use of improved cultivars with N applications timed to coincide with maximum crop demand at 45 and 70 DAT, could enhance N fertilizer use efficiency for increased rice yields as well as reduce N losses downstream, which can cause adverse off-site environmental effects.