9-Acridone-4-carboxylic acid has been established as an efficient Cr(III) fluorescent sensor. The binding of this ligand with Cr(III) is confirmed by FTIR, thermal and mass spectral analysis of the product. Based on this chelation assisted fluorescence quenching, a highly sensitive spectrofluorometric method is developed for trace level detection, estimation and speciation studies of chromium in DMF-water. The ligand has an excitation and emission maxima at 408nm and 498.4nm, respectively. The equilibrium binding constant of the ligand with Cr(III) is 8.1378×10 4 as calculated using Stern–Volmer equation. Up to 9×10 −6 molL −1 of [Cr 3+ ], linearity has been observed. The interference of foreign ions has been found to be negligible.