AbstractDenitrification is a key process in the global nitrogen (N) cycle, causing both nitrous oxide (N2O) and dinitrogen (N2) emissions. However, estimates of seasonal denitrification losses (N2O + N2) are scarce, reflecting methodological difficulties in measuring soil‐borne N2 emissions against the high atmospheric N2 background and challenges regarding their spatio‐temporal upscaling. This study investigated N2O + N2 losses in response to N fertilizer rates (0, 100, 150, 200, and 250 kg N ha−1) on two intensively managed tropical sugarcane farms in Australia, by combining automated N2O monitoring, in situ N2 and N2O measurements using the 15N gas flux method and fertilizer 15N recoveries at harvest. Dynamic changes in the N2O/(N2O + N2) ratio (<0.01 to 0.768) were explained by fitting generalized additive mixed models (GAMMs) with soil factors to upscale high temporal‐resolution N2O data to daily N2 emissions over the season. Cumulative N2O + N2 losses ranged from 12 to 87 kg N ha−1, increasing non‐linearly with increasing N fertilizer rates. Emissions of N2O + N2 accounted for 31%–78% of fertilizer 15N losses and were dominated by environmentally benign N2 emissions. The contribution of denitrification to N fertilizer loss decreased with increasing N rates, suggesting increasing significance of other N loss pathways including leaching and runoff at higher N rates. This study delivers a blueprint approach to extrapolate denitrification measurements at both temporal and spatial scales, which can be applied in fertilized agroecosystems. Robust estimates of denitrification losses determined using this method will help to improve cropping system modeling approaches, advancing our understanding of the N cycle across scales.