Abstract: A highly sensitive and selective spectrophotometric method was developed for the determination of trace amount of uranium (VI) in water samples in aqueous DMF medium. The uranyl ion forms a pale yellow colored complex with 5-Bromo salicylaldehyde isonocotinoyl hydrazone (5-BrSAINH) in an acidic buffer of pH 5.0. The complex has an absorbance maximum at 395nm, was stable for more than 72 hours. The complex shows maximum absorbance at 395nm. Beer’s law is obeyed in the range 0.119-1.071 µgml-1. The molar absorptivity and sandell’s sensitivity of the proposed method were found as 1.17 x 104 L.mol-1cm-1 and 0.02µgcm-2,respectively.The pale yellow colored complex has 1:1 (Uranium (VI)- 5BrSAINH) Stoichiometry .The stability constant of the complex was determined as 1.80x105 by Job’s method. A sensitive and selective second order derivative spectrophotometry has also been proposed for the determination of U (VI). The interference of various cations and anions were studied. The developed methods were successfully employed in the determination of U (VI) in environmental, phosphate rocks and fertilizer samples.