The aim of the study was to evaluate the effect of nitrogen (N) fertilisation on certain quality parameters and in vitro total gas and methane production of improved grass species commonly used as fodder species in South Africa. Treatments included seven grass species representing two photosynthetic pathways (C3 and C4) with three levels of N fertilisation (0, 50 and 100 kg N ha–1). Plants were grown in a greenhouse and N was applied in a single application after a simulated defoliation. Sample material was harvested by hand after an 8-week regrowth period. Grass species and rate of N fertiliser both had effects (P < 0.05) on the nutritive value and in vitro organic matter digestibility of the selected species. Crude protein concentration increased (P < 0.05) and neutral detergent fibre concentration tended to decrease as the level of N fertilisation increased for both C3 and C4 species. Generally, no effect was found of N fertilisation on in vitro total gas or methane production; however, increasing the level of N fertiliser increased (P < 0.05) the methanogenic potential (in vitro methane/in vitro total gas production) of D. glomerata, F. arundinacea and C. ciliaris after a 24-h incubation period but no significant effects were reported after a 48-h incubation period.