A differential spectrophotometric method has been developed for uranium in presence of plutonium by making absorbance measurements at 420 nm in 4M H2SO4 using 5 cm cells. The absorbance measurements are made with two independent sets of standards: (1) having uranium only and (2) having uranium and plutonium in a fixed ratio R, against a uranium solution of high absorbance (1A) in the reference beam. A least-squares fit of data on absorbance and uranium concentration in the two cases gave two slopes m1 and m2, which were used to determine the concentration of uranium using the relationship CU=C0+m1·[AT-(1/m2–1/m1) R·CPu] where AT is the relative absorbance of uranium and plutonium at 420 nm and C0 is the intercept corresponding to slope m1 for pure uranium standards and m2 is the slope for mixed uranium and plutonium standards. A knowledge of CPu, the plutonium concentration, is essential and is obtained by differential spectrophotometric measurements at 835 nm by oxidizing plutonium to its hexavalent state. In the same aliquot, plutonium could be determined with a precision of better than ±0.5% and uranium with a precision of better than ±1.0%.