Persistence of dissolved organic matter (DOM) in aquatic environments may in part be explained by high diversity and low concentrations of carbon substrates. However, changes in dissolved substrate quality can modify aquatic bacterial community composition and rate of carbon uptake. The aim of this study was to test if the presence of multiple simple substrates affects the turnover of organic carbon. Natural bacterial communities were grown in continuous cultures supplied with either individual carbon substrates—salicylic acid (SA), tryptophan (Trp) or tyrosine (Tyr)—or a combination of the 3 substrates. Concentrations were tracked using fluorescence spectroscopy, and steady-state concentrations of a few nanomolar were reached. Bacterial growth efficiency was dependent on which carbon sources were present and reached an intermediate level in the combined treatment. The bacterial community maintained steady-state concentrations of Trp that were lower in the combined treatment than in the individual substrate treatment. In addition, steady-state concentrations were reached faster during growth on combined carbon substrates, although the maximum utilization rate of each individual compound was lower. However, the steady-state concentration of total carbon (sum of carbon content of SA, Trp and Tyr) was higher in the combined culture than in the individual substrate treatments, and seemed to be determined by the carbon substate for which the bacteria had the lowest affinity. The results from this study indicate that persistence of dissolved organic carbon can in part be explained by vast substrate diversity, which raises the threshold concentration for utilization by natural bacterial communities.