Amino acid and oligopeptide-nitrogen (N) forms only a minor component of the total dissolved N pool in grassland soils, yet the importance of these N-pools for plant productivity will ultimately depend on the rate at which these pools turnover. Fluxes of dissolved organic matter (DOM) through the soil solution are frequently estimated from measurements of respiration, but this method fails to consider any delay between microbial substrate acquisition and mineralization. Here, we added 14C-labelled alanine and tri-alanine (10μM) to 4 soils collected from a natural grassland productivity gradient and then measured substrate depletion from the soil solution and the subsequent production of 14CO2 resulting from mineralization at 1-60min. There was a considerable delay between microbial 14C removal from the soil solution, which occurred extremely rapidly (up to 96% of added substrate depleted within a minute), and 14CO2 evolution resulting from the fast turnover of the alanine and tri-alanine. This indicates that amino acid and peptide longevity in the soil solution of the soils in this grassland productivity gradient has been greatly overestimated from measurements of mineralization alone. Rates of substrate uptake and mineralization by microbes declined in less productive, N-limited grassland soils with lower levels of microbial biomass, suggesting that the availability of organic N for plant uptake is likely to be controlled by soil microbial activity. We estimate that amino acid and peptide pools occurring in the most productive grassland soils may turnover at a rate of up to 20 times a minute, representing a very considerable flux of N through the soil.