The relationships between leaf structure, nitrogen concentration and CO2 assimilation rate (A) were studied for 14 grass species grown in the laboratory under non-limiting nutrient conditions. Structural features included leaf thickness and density, and the proportion of leaf volume occupied by different types of tissue (mesophyll, epidermis, vessels and sclerenchyma). Relationships were assessed for data expressed per unit leaf area and fresh mass. The latter was found to be closely related to leaf volume, which allowed us to use A per unit leaf fresh mass (Afm) as a surrogate of A per unit leaf volume. Assimilation rate per unit leaf area (Aa) was positively correlated with leaf thickness and with the amount of mesophyll per unit leaf area; the relationship with leaf nitrogen content per unit area was only marginally significant. Afm was negatively correlated with leaf thickness and positively with fresh mass-based leaf organic nitrogen concentration. A multiple regression involving these two variables explained 81% of the variance in Afm. The value of Afm was also significantly related to the proportion of mesophyll in the leaf volume, but surprisingly the correlation was negative. This was because thin leaves with high Afm and nitrogen concentration had proportionally more mechanically supportive tissues than thick ones; as a consequence, they also had a lower proportion of mesophyll. These data suggest that, in addition to leaf nitrogen, leaf thickness has a strong impact on CO2 assimilation rate for the grass species studied.