Abstract Purpose Large parts of the Amazon rainforest grow on weathered soils depleted in phosphorus and rock-derived cations. We tested the hypothesis that in this ecosystem, fine roots stimulate decomposition and nutrient release from leaf litter biochemically by releasing enzymes, and by exuding labile carbon stimulating microbial decomposers. Methods We monitored leaf litter decomposition in a Central Amazon tropical rainforest, where fine roots were either present or excluded, over 188 days and added labile carbon substrates (glucose and citric acid) in a fully factorial design. We tracked litter mass loss, remaining carbon, nitrogen, phosphorus and cation concentrations, extracellular enzyme activity and microbial carbon and nutrient concentrations. Results Fine root presence did not affect litter mass loss but significantly increased the loss of phosphorus and cations from leaf litter. In the presence of fine roots, acid phosphatase activity was 43.2% higher, while neither microbial stoichiometry, nor extracellular enzyme activities targeting carbon- and nitrogen-containing compounds changed. Glucose additions increased phosphorus loss from litter when fine roots were present, and enhanced phosphatase activity in root exclusions. Citric acid additions reduced litter mass loss, microbial biomass nitrogen and phosphorus, regardless of fine root presence or exclusion. Conclusions We conclude that plant roots release significant amounts of acid phosphatases into the litter layer and mobilize phosphorus without affecting litter mass loss. Our results further indicate that added labile carbon inputs (i.e. glucose) can stimulate acid phosphatase production by microbial decomposers, highlighting the potential importance of plant-microbial feedbacks in tropical forest ecosystems.