Vertical distribution (0-15cm) of the macrobenthic community and its relationships to natural sediment characteristics and trace metal contents and bioavailability were studied at five locations in the lower Douro estuary, Portugal. An analysis of vertical metal distribution, for the interpretation of anthropogenic impact on the estuarine sediments, was also investigated. Sediment characterisation included organic matter, grain size, metals (Al, Fe, Cu, Pb, Cr, Ni, Cd, Zn and Mn), acid volatile sulphide (AVS) and simultaneously extracted metals (SEM). The macrobenthic community had low diversity (14 species), was dominated by small size opportunists and seemed to be controlled mainly by natural factors such as grain size distribution, Al and Fe contents and sediment depth. The vertically heterogeneous distribution of macrobenthic community appears to affect redox status of the sediments and consequently metal bioavailability. Despite anthropogenic contamination in terms of Zn, Cu, Pb, Cr and Ni having already been detected in the north bank, the analysis of vertical distribution was essential for the identification of current anthropogenic contamination in terms of Zn, Pb and Cd in the south bank.