Abstract: A method of FI-FAAS for simultaneous determination of Cr(Ⅵ) and Cr(Ⅲ) with on-line separation of Cr(Ⅵ) from Cr(Ⅲ) by adsorption on TBP-cotton fiber was proposed. A self-made conical column was used to pack the TBP-cotton fiber globes, and incorporated into the FI system as an on-line separation unit. The sample solution with its acidity at pH 0.75 was injected into the FI-system at the flow-rate of 4 mL·min-1 and separated (or enriched) in the conical column for 160 s, having Cr(Ⅵ) adsorbed on TBP-cotton fiber, and Cr(Ⅲ) flowed out in the effluent which was collected and used for determination of Cr(Ⅲ). Cr(Ⅵ) adsorbed on the column was eluted with distilled water with a flow-rate of 2.6 mL·min-1, and the eluate was introduced into FAAS for determination of Cr(Ⅵ). Linear relationship between values of absorbance and mass concentration of chromium was kept in the range of 0.100-0.900 mg·L-1. Precision was tested by repetitive determinations of 0.10 μg Cr(Ⅵ) in presence of 0.02 μg Cr(Ⅲ), and value of RSD (n=7) found for Cr(Ⅵ) was 6.4%. Recovery was tested by analyzing mixed solutions containing 0.500 mg·L-1 Cr(Ⅵ) and 0.100 mg·L-1 Cr(Ⅲ), giving average recovery of 119% for Cr(Ⅵ), and of 107% for Cr(Ⅲ).