The research was carried out in a Mediterranean semi-arid environment on berseem clover, annual ryegrass and their mixture to study the effect of defoliation management [date of first cut (FC) 85, 119, 140, 169 days after sowing] and different plant arrangements (sowing the two components in alternate rows or in the same row) on yields, N content, N2 fixation and N transfer. The experimental design was a split-plot with four replications. The 15 N isotope dilution technique was used (8 kg N ha –1 as ammonium sulphate at 10 atom% 15 N excess) to evaluate the N2 fixation. Total seasonal DM yield was, on average, significantly higher for FC119 and FC140 (approx. 12.3 t ha –1 ) than for FC85 and FC169 (approx. 10.6 t ha –1 ). Plant arrangement did not significantly influence total yield of the mixture. However, the legume yield was higher (+20%; P<0.0001) in the same row than in alternate rows arrangement. N content of ryegrass was significantly higher in the mixtures than in pure stand and in the ‘same row’ plant arrangement than in the ‘alternate rows’. Intercropped berseem always had a significant higher % of Ndfa than the monocropped one (on average 74.7% and 57.7% respectively). The apparent transfer of fixed N from berseem to ryegrass was not detected in either plant arrangement.