Headwater forest ecosystems of the western USA generate a large portion of the dissolved organic matter (DOM) transported in streams across North America. Land cover changes that alter forest structure and species composition affect the quantity and composition of DOM transferred to aquatic ecosystems. Clear-cut harvesting affects ~1% of the forest area of North America annually, leaving most western forests in varying stages of regrowth and the total area of old-growth forest is decreasing. The consequences of this widespread management practice on watershed carbon cycling remain unknown. We investigated the role of land cover change, because of clear-cut harvesting, from mixed-species old-growth to lodgepole pine-dominated second-growth forest on the character and reactivity of hillslope DOM exports. We evaluated inputs of DOM from litter leachates and export of DOM collected at the base of trenched hillslopes during a 3-year period (2016–2018) at the Fraser Experimental Forest in north-central Colorado, USA. Dissolved organic carbon and total dissolved nitrogen were higher in lateral subsurface flow draining old- versus second-growth forest. Fluorescence spectroscopy showed that the DOM exported from the old-growth forest was more heterogeneous and aromatic and that proteinaceous, microbially processed DOM components were more prevalent in the second-growth forest. Biological oxygen demand assays revealed much lower microbial metabolism of DOM in litter leachate and subsurface exports from the old-growth forest relative to second growth. Old-growth and second-growth forests are co-mingled in managed ecosystems, and our findings demonstrate that land cover change from a mixture of conifer species to lodgepole pine dominance influences DOM inputs that can increase the reactivity of DOM transferred from terrestrial to aquatic ecosystems.