Nitrogen (N) fertilization is important for sugarcane crops, for both agronomic and environmental reasons. Better use of N from fertilizer is required for crop management, and in order to achieve it, adequate N nutrition monitoring is necessary. Sap extract analysis is a sensitive technique that has been adopted in other crops to tackle this objective, and can be used for sugarcane. The aim of this study was to evaluate the behavior of N and the response of sugarcane after N fertilization, using sap and leaf tissue analysis. The study was undertaken in two experimental areas in 2012/2013 season, using ratoon cane, following application of 0, 50, 100, and 150 kg•ha −1 of N. The youngest visible leaves were sampled from sugarcane at five time points between 0 and 150 days after fertilization, to evaluate the nitrate, ammonium, and total N content of sap extract, and total N content of leaf tissue. At the first sampling point (30 days after N fertilizer application), mineral N in sap increased as a result of N fertilization in medium-textured soil, and remained high until 120 days after fertilization in clayey soil. The total N content of sap followed a similar behavior. Stalk yield was only observed to increase in sugarcane grown in the clayey soil. Sap analysis proved to be an efficient method for monitoring N status in sugarcane grown in different soil types.