An accurate assessment of soil organic matter dynamics is critical for understanding and predicting ecosystem responses to anthropogenic perturbation such as climate change, pollution and agriculture. These measurements of carbon (C) turnover are frequently made in the laboratory where it is easier to control the experimental conditions. However, this could potentially bias the results due to a range of factors including the absence of plants and associated mycorrhizas, physical disturbance, alteration in redox status etc. The aim of this study was therefore to directly compare the turnover of organic acids under field and laboratory conditions in three contrasting land use regimes (grassland, woodland and arable). The method involved the injection of 14C-labeled citric acid into the top soil and monitoring of its mineralization by capturing 14CO2 evolved from the soil over a 7 day period. Laboratory climate conditions were matched to those in the field. In both the laboratory and the field we showed that 14CO2 evolution followed a bi-phasic pattern and conformed extremely well to a double first-order kinetic model. While the first rapid mineralization phase showed a similar half-life for citrate under both laboratory and field conditions (4.9 ± 0.7 h), the second slower mineralization phase had a significantly longer half-life when performed in the field. Overall, our results suggest that the first rapid mineralization phase was largely independent of our experimental conditions whilst the turnover of the citrate-derived C immobilized in the microbial biomass was significantly affected by the incubation conditions either due to differences in substrate utilization and storage pathways or due to faster microbial turnover in the laboratory.