Ammonium sulfate, labeled with the stable N-15 isotope, was applied to the soil at the rate of 45 kg/ha of actual nitrogen to 7-year-old 'Royal Ann' trees on 'Mazzard' rootstock. The fertilizer was applied at four timings: rapid shoot growth/spring (10 May), pre-harvest (15 June), post-harvest (3 Aug) and pre-leaf fall (3 Oct) to eight replicate trees. Four trees were excavated at the end of the growing season, prior to leaf fall and another set of four after a second growing season. Fruit were harvested in mid-June of both years. Trees were excavated and partitioned into fruit, current season shoot growth, numerous wood components, spurs, roots and leaves. Components were sub-sampled, dried and analyzed for total N and N-15 content. In October, whole tree total N was 2.4, 2.4, 1.9 and 2.0 kg per tree for spring, pre-harvest, post-harvest and pre-leaf fall applications, respectively. The respective amounts of fertilizer N was 0.30, 0.16, 0.09 and 0.06 kg/tree. N derived from the fertilizer (NDFF) declined from 13% to 3% and uptake efficiency also declined from 21% to 4% for the different timings. After the second growing season, NDFF and uptake efficiency were not different between the timings and had a mean of 14% and 26%, respectively.