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Respiration of dissolved organic matter (DOM) in streams contributes to the global CO₂ efflux, yet this efflux has not been linked to specific DOM sources and their respective uptake rates. Further, removal of DOM inferred from longitudinal concentration gradients in river networks has been insufficient to account for observed CO₂ outgassing. We hypothesize that understanding in‐stream dynamics of DOM, which is a heterogeneous mixture spanning a wide range of biological labilities, requires considering that DOM lability decreases during downstream transport. To test this hypothesis, we paired seasonal bioreactor measurements of DOM biological lability with whole‐stream tracer data from White Clay Creek, Pennsylvania, USA, and used a particle‐tracking model to predict in‐stream DOM dynamics. The model simulates continuous inputs of DOM and uses storage time in the stream bioactive regions plus kinetic parameters from bioreactors to assess differential uptake of DOM fractions (i.e. fractionation) in the stream. We compared predictions for in‐stream dynamics of bulk DOM concentration (quantified as dissolved organic carbon) and fluorescent DOM components. Our model‐data synthesis approach demonstrates that more labile fractions of DOM in streamwater preferentially originate and are consumed within short travel distances, causing spiraling metrics to change with downstream distance. Our model can account for local sources of rapidly‐cycled labile DOM, providing a basis for improved interpretation of DOM dynamics in streams that can reconcile apparent discrepancies between respiratory outgassing of CO₂ and longitudinal DOM concentration gradients within river networks.

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