Understanding the interactions between the initial biochemical composition and subsequent decomposition of plant litter will improve our understanding of its influence on microbial substrate use to explain the flow of organic matter between soil carbon pools. We determined the effects of land use (cultivation/native woodland/native pasture), litter type (above and below ground) and their interaction on the initial biochemical composition (carbon, nitrogen, water soluble carbon, lignin, tannin and cellulose) and decomposition of litter. Litter decomposition was studied as the mineralization of C from litter by microbial respiration and was measured as CO₂-C production during 105 d of laboratory incubation with soil. A two-pool model was used to quantify C mineralization kinetics. For all litter types, the active C pool decay rate constants ranged from 0.072 d⁻¹ to 0.805 d⁻¹ which represented relatively short half-lives of between 1 and 10 days, implying that this pool contained compounds that were rapidly mineralized by microbes during the initial stages of incubation. Conversely, the decay rate constants for the slow C pool varied widely between litter types within and among land uses ranging from 0.002 d⁻¹and 0.019 d⁻¹ representing half-lives of between 37 and 446 days. In all litter types, the initial lignin:N ratio strongly and negatively influenced the decay rate of the slow C pool which implied that the interaction between these two litter quality variables had important controls over the decomposition of the litter slow C pool. We interpret our results to suggest that where the flow of C from the active pool to the slow pool is largely driven by microbial activity in soil, the rate of transfer of C will be largely controlled by the quality of litter under different land-use systems and particularly the initial lignin:N ratio of the litter. Compared with native pastures and cultivation, above and below ground litter from native woodland was characterized by higher lignin:N ratio and more slowly decomposing slow C pools which implies that litter is likely to persist in soils, however based on the sandy nature of the soils in this study, it is likely to lack protection from microbial degradation in the long term.