Nitrogen (N) partitioning between trees and crops in alley-cropping systems (AC) may increase the N fertilizer use, but more investigation is needed to elucidate it for young trees. The aim of this study was to explore the share of N between crops and three tree species after a few years of tree establishment in AC. Using 15N labeling (10 atm% as 15NH4,15NO3 form), we traced the fate of N fertilizer in wheat associated with four-year-old trees at Ramecourt alley-cropping experimental site. The 15N abundance of tree leaves and crop whole-plant was compared between samples collected in AC, in sole-crop control (CC) and in unlabeled and unfertilized pure-forest control plots (FC). The wheat δ15N increased from 0.7 ± 0.6‰ to 994 ± 474‰ before and 12 days after labeling and remained stable until harvest. For natural abundance, no significant difference in wheat δ15N was observed between AC and CC. The same result was observed for labeled treatment. The percentage of wheat N derived from fertilizer was on average 27 ± 17% and no effect of type of system or associated tree species was observed. Tree foliar δ15N was multiplied by 1.5, 1.8 and 4 for willow, hornbeam and wild cherry respectively compared to unlabeled trees in AC. The percentage of tree foliar N derived from fertilizer was on average 0.04 ± 0.04% and no significant difference was observed between tree species. The wheat associated with trees allocated significantly more tracer N to ears than wheat in monocrop. From this study, we succeeded in quantifying the beginning of tree/crop sharing of N fertilizer using labeling method and evidenced tree species effect. Young trees in AC contributed to valorize the N fertilizer without significant effect on global wheat uptake and favored its valorization in wheat grain.