Abstract: As a novel salt-free reductant in U/Pu separation and Pu purification process, hydroxyl-semicarbazide (HSC) can effectively adjust the valence of plutonium in reprocessing solutions, and its concentration is a vital parameter and necessary to be determined accurately. Under the catalysis of acid, color reaction between HSC and paradimethylaminobenzaldehyde can occur, which makes it possible to determine the concentration of HSC by spectrophotometry. Experimental results show that concentrations of HSC within 2.5-60.0 μmol/L with its absorptions conform to Beer law. The linear correlation coefficient R² is 0.999 9, the measuring precision of better than 5.0%, and the recoveries of actual samples are 95%-105%. Experimental results also show that the standard curves are good repeatability and conformity in 45 days.