The speciation of thallium was investigated in the Indian mustard plant (Sinapis alba L.), hydroponically cultivated and in moss (Entondon schreberi) used as a bioindicator. The separation of thallium species was done using anion exchange chromatography coupled with on-line ICP-MS detection with 100 mmol L⁻¹ ammonium acetate containing 5 mmol L⁻¹ DTPA as a mobile phase. The study indicated that Tl(I) is the dominating form in plant extracts; Tl(III) was not detected. Three species of Tl were found in the leaves, two of them remained unidentified. Additionally, the fractionation study of Tl in organs of the Indian mustard plant using water, ammonium acetate containing DTPA and sodium dodecyl sulfate solutions was performed. The highest leaching efficiency was reached using an ammonium acetate/DTPA solution. Next, the efficiency of Tl transport from roots to stems and leaves and the accumulation factors were estimated. 80% of uptaken Tl was found in above-ground plant organs.