Forest-to-pasture conversion has been reported to increase soil organic matter (SOM) in mineral topsoils in the tropical mountain rainforest region of south Ecuador, with subsequent decreases following pasture abandonment. Until now the mechanisms behind these changes have not been fully understood. To elucidate their varied preservation patterns, we analysed root- and shoot-derived organic matter and assessed their contribution to the formation of SOM in topsoils (0–5 cm) on a chronosequence of pastures (Setaria sphacelata (Schumach.); C4) established after slash and burn of the natural forest (diverse C3 plant species) and an abandoned pasture site invaded by bracken fern (Pteridium arachnoideum (Kaulf.) Maxon.; C3). Cutin and suberin biomarkers of the two plant species (grass and bracken) and of forest litter were identified after saponification and their contribution to SOM was studied by compound-specific stable carbon isotope analyses. Our results showed specific root and shoot biomarkers for the two plant species and for forest litter, which often did not correspond to the classification of root-versus shoot-specific monomers reported in the literature. This illustrates the importance of direct biomarker determination rather than using results from studies with different plants. Shoot- as well as root-derived OM of forest and grass origin contributed to the stable SOM pool with decadal turnover times. Forest-derived monomers contributed more to the stable SOM pool compared with grass-derived monomers. ω-hydroxy carboxylic acids and α,ω-alkanedioic acids of forest origin may have been stabilized in these tropical soils by bonding to soil minerals. Rapid degradation of grass-derived lipids from the same compound classes suggests a saturation of the mineral binding capacity. In pasture soils, the accumulation of SOM was mainly driven by large inputs of root OM. The accumulated SOM during pasture use is, however, lost rapidly after abandonment.