Background and aims: Plant-derived, ester-bound substituted fatty acids have been used for decades as biomarkers to identify input of plant materials in sediments and soils. However, the long-term decomposition patterns of these biomarker compounds under natural conditions are not well understood, although this is a basic prerequisite for quantitative biomarker applications. Methods: For this study, we analyzed the decomposition patterns of root- and needle-specific compounds of Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) in a litterbag study conducted over 3 years. Samples were analyzed by methanolic KOH extraction with previous removal of free extractable lipids. Results: The concentrations of most detectable compounds had decreased after three years of incubation. The observed changes of concentrations followed a non-linear path and cannot be explained by microbial uptake and metabolism alone. Other factors controlling the breakdown of ester-bound lipids, like lipid oxidation must play a role. Between similar plant parts and different plant parts of the same species, the observed degradation patterns were heterogeneous. The estimated ratio of remaining root and needle biomass that may arise with the choice of a particular biomarker varies in this study between 0.6 and 40 times after three years. Conclusion: This range of variation does not allow reliable conclusions about the contribution of roots and needles to decomposed organic matter based on biomarkers ratios.