In Brazil, N fertilisers are applied onto the surface of the sugarcane (Saccharum spp.) litter layer, which represents a large portion of the C stored in the agroecosystem. However, little is known about the influence of mineral N on decomposition of organic C compounds and mineral N transformations in the litter-soil transition zone. This investigation determined the effect of mineral N forms (NH4+ and NO3-) on glucose mineralisation and N dynamics in litter and topsoil from two sugarcane fields located in the state of São Paulo, Brazil. Carbon mineralisation and N availability were measured using 14C-labelled glucose, applied alone (glucose) or combined with NH4+ [as (NH4)2SO4] (glucose + ammonium) or NO3- (as KNO3) (glucose + nitrate) in litter and soil from the litter-soil transition zone. The 14C immobilised after 168 h had the following trend for the litter of both sites: glucose > glucose + ammonium > glucose + nitrate, while no differences among treatments were found for the soils. Higher immobilisation (increased by 165%, on average) occurred for NH4+ (glucose + ammonium) compared to NO3- (glucose + nitrate), primarily in the sugarcane litter. Carbon mineralisation in litter was N-limited, and the addition of mineral N accelerated the return of C to the atmosphere. Lower immobilisation of NO3- may increase N availability to plants relative to NH4+ in the short term, but also may lead to reduced C storage and larger greenhouse gas emissions due to faster mineralisation of labile carbohydrate through the NO3- reduction process.