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Abstract The Río Tinto mining area provides ideal conditions for studying the role of colloidal particles in concentrating. All the elements are present in higher concentrations respect to their content in common natural waters. 234U/238U activity ratio ranges between 2.286 ± 0.149 and 2.531 ± 0.151. Concentration of Fe and Al are in the order of 104–103 mg/L; Co, Zn and Cu values are three times lower than Fe and Al, but still much greater than in natural waters; U reaches values up to 40.73 μg/L. To evaluate the partitioning of these elements into the colloidal fraction, ultrafiltration (UF) experiments were made. A permeation model describing the relationship between concentration of the elements or isotopes in the permeate solution and the concentration factor (CF) was calculated. Experiments were carried out using CF from 1.5 to 25.0. The behaviour of investigated heavy metals and U can be predicted by a permeation model that provides similar permeation coefficient (P c) for all of them (P c = 0.95–0.97 for 50 kDa, P c = 0.86–0.88 for 10 kDa, and P c = 0.80–0.86 for 3 kDa). Retention for all the investigated elements occurs, resulting in up to 23% of the element associated to the 3 kDa colloidal fraction. The increase in U and heavy metals concentrations in the retentate fraction with the increasing CF is due to the retention of the colloidal fraction. The colloidal abundance of related metals increased with decreasing membrane MWCO even though the difference is small between 3 and 10 kDa fraction for some metals Characterization of colloidal material is needed to better understand the state of play in this context. Highlights • Partitioning of U and selected heavy metals in dissolved and colloidal fractions in AMD conditions were examined. • UF with distinct MWCO was used and a permeation model to quantify the behaviour of U and selected metals was applied. • Metal concentrations much higher than those in natural waters were found and high 234U/238U in the water samples were observed. • Little colloidal metals in the size fraction 3–10 kDa are present, likely due to the strong acidic conditions. • Low concentrations of colloidal U in the study area were measured. [ABSTRACT FROM AUTHOR]