The quality of grass in terms of form and relative amounts of energy and protein affects both animal production per unit of intake and nitrogen (N) utilization. Quality can be manipulated by herbage management and choice of cultivar. The effects of N application rate (0, 90 or 390 kg N ha−1 year−1), duration of regrowth period (2–3, 4–5, or 6–7 weeks), and cutting height (8 or 12 cm) on the mass fractions of nitrogen (N), water-soluble carbohydrates (WSC), neutral detergent fibre (NDF), acid detergent fibre (ADF), lignin and ash in lamina and sheath material of a high-sugar (Aberdart) and a low-sugar (Respect) perennial ryegrass (Lolium perenne) cultivar, were studied in a factorial field experiment during four seasons in 2002 and 2003. Expressing NDF and ADF mass fractions in g per kg WSC-free dry matter (DM) increased the consistency of treatment effects. The high-sugar cultivar had generally higher WSC mass fractions than the low-sugar cultivar, especially during the late season. Moreover, the relative difference in WSC mass fraction between the two cultivars tended to be higher for the lamina material than for the sheath material, which suggests that the high-sugar trait may be more important under grazing conditions, when lamina forms the bulk of the intake, than under mowing regimes. Longer regrowth periods and lower N application rates increased WSC mass fractions and decreased N mass fractions; interactions between regrowth period and N application rate were highly significant. The mass fractions of NDF and ADF were much less influenced. The NDF mass fraction in terms of g per kg WSC-free DM tended to be higher at lower N application rates and at longer regrowth periods. The effect of cutting height on herbage chemical composition was unclear. In conclusion, high-sugar cultivars, N application rate and length of the regrowth period are important tools for manipulating herbage quality