Abstract In present research, a comparative validation study for the estimation of cefotaxime sodium in pharmaceutical formulations by spectrophotometric and fluorimetric methods were performed. The developed methods depend on the determination of cefotaxime sodium by using 2, 4-Dinitro phenyl hydrazine. By using 2, 4-Dinitro phenyl hydrazine, these drug form ion-pair complex in acidic medium. This coloured complex shows maximum absorbance at 408nm by spectrophotometric method (Method A) and fluorescent intensity was measured by using fluorimetric method (Method B).Linearity calibration curve were obtained in a concentration range of 1.0-6.0 µg/ml for cefotaxime sodium by using both of these methods. The result of analysis of these methods shows that the amount of drugs present in the formulation has a very good correlation with the label claim of the formulation. The developed methods were validated for various parameters as per ICH guidelines like Accuracy, Precision, Linearity, and range, LOD and LOQ, Ruggedness and Robustness. %RSD will be less than 2 for all the validation parameters. Recoveries studies revealed that results within the specified limits. Hence the proposed spectrophotometric and fluorimetric methods were found to be satisfactory and could be used for the routine analysis of cefotaxime sodium in their marketed formulations. Statistical comparison of the results with the reference methods showed excellent agreement and proved that no significant difference in the accuracy and precision. Keywords: Cefotaxime sodium; 2, 4-Dinitrophenyl hydrazine; Fluorimeter and UV-Visible Spectrophotometer.