A quantification procedure of trace elements during colloid size fractionation was developed and validated. This procedure is based on the hyphenation between Asymmetrical Flow Field-Flow Fractionation (As-Fl-FFF) and Inductively Coupled Plasma Mass Spectrometer (ICP-MS). The optimisation of the procedure was performed on a soil leachate spiked with six trace elements selected for their environmental and health impact (As, Cd, Sb, Se, Sn and Pb). The elements in the spiked sample were on-line monitored during the fractionation. The validation was carried out by comparison with a second off-line quantification procedure based on fraction collection and total element analysis by ICP-MS. This off-line one was previously validated using reference materials. Finally, the analytical performances of the two procedures were compared.