Hydride generation reactions are frequently used for the speciation analysis of organotin compounds in a wide variety of matrices. However, the organotin determination procedure may be altered by different interferences mechanisms. This work evaluates the interferences likely to occur in the determination of tributyltin in a harbour sediment candidate reference material for the EEC Community Bureau of Reference with the method of hydride generation combined on-line with cryogenic trapping, gas chromatographic separation and detection by atomic absorption spectrometry. First, a full geochemical characterization of the sediment and of its acetic acid leachate was achieved to evaluate interfering compounds that may inhibit the determination of TBT by hydride generation in this sediment. Second, interference studies on various organic and inorganic compounds likely to alter the overall organotin determination were carried out with simple model solutions. These model solutions were spiked successively with different possible interfering compounds mixed with the organotins under study, i.e., monomethyltin (MMT), monobutyltin (MBT), dibutyltin (DBT) and tributyltin (TBT) in a mixed solution at concentrations of 5 ng as Sn of each compound in 50 ml of water. Organic interferents studied were organic pollutants (organic solvents, polychlorinated biphenyls, pesticides, n-alkanes), humic substances and a complexing agent, EDTA. The inorganic interferents were represented by a mixture of fourteen inorganic metals. The addition of organic compounds to the hydride generation reaction had little effect on the signal suppression for all organotin species. Reproducibility was significantly affected only by the presence of humic substances in solution. Signal suppression was observed only for the monoalkyltins (MMT and MBT) in the presence of EDTA. Solutions of mixed inorganic metals were found to be highly efficient in inducing severe signal suppression for all organotin compounds at a level of 100 μg of each element. Interference mechanisms during the speciation determination step are outlined and discussed.