Abstract An aqueous two-phase system of tetrabutylammonium bromide (TBAB) and (NH 4) 2SO 4 mixture has been developed for the extractive preconcentration and separation of certain compounds. TBAB concentration in upper phase is much higher than that of bottom solution. This new aqueous two-phase system is proposed for the highly selective extraction of Cr(VI) from large amounts of Cr 3+. The Cr(VI) is found to be extracted into the TBAB-rich upper phase due to ion pair formation such as HCrO 4 − · TBAB +. The Cr(VI) was sufficiently extracted into the upper phase in the pH range from 1 to 5. The proposed method has been applied to the determination of trace of Cr(VI) in wastewater samples with a coefficient of variation less than 3.2%. The recovery obtained was not lower than 90%. The determination limit for the Cr(VI) was found to be 60 μg l −1 in 10 ml of sample solution.