A highly sensitive, selective, and rapid method for the determination of ng mL level of U(VI) based on the rapid reaction of U(VI) with 2-(2-benzothiazolylazo)-3-hydroxyphenol (BTAHP) and the solid-phase extraction of the colored complex with a reversed-phase polymer-based C18 cartridge was developed. The BTAHP reacted with U(VI) to form a violet complex of molar ratio 2:1 [BTAHP to U(VI)] in the presence of 4.0 M of phosphoric acid solution and Triton X-114 medium. This complex was enriched by the solid-phase extraction with a polymer-based C18 cartridge. The enrichment factor of 200 was achieved. The molar absorptivity of the complex is 2.73 × 10 L mol cm at 639 nm in the measured solution. The system obeys Beer's law in the range of 2.0–125 ng mL, whereas the optimum concentration range obtained from Ringbom plot was 8.0–115 ng mL. The relative standard deviation for 10-replicates sample of 100 ng mL level is 1.05%. The detection and quantification limits are 0.6 and 1.98 ng mL in the original sample. This method was applied to the determination of uranium(VI) in sea, tap, and waste waters, ore standard reference material, soil and sediment samples with good results comparing to the graphite furnace atomic absorption spectroscopy (GFAAS) method.