An oxygen insensitive azoreductase was purified from a novel bacterial strain (Staphylococcus sp. KU898286) that was isolated from an abandoned site of the textile waste discharge unit. The isolated enzyme had efficiently cleaved the azo-bonds through reductive transformation under aerobic conditions. Initial phenotypic characterization and final construction of phylogenetic tree on the basis of 16s rDNA demonstrated 99% resemblance of the isolate to Staphylococcus aureus. The purified azoreductase was found to have a broad spectrum activity that reduced RR241 at a concentration of 50mg/L with pH between 6–8 and 30°C temperature). Besides, the reactive red 241 (RR241) was reduced at extracellular level as well as NADH dependent intracellular level. Complete reduction/ decolourization of RR241 were achieved after 18 hrs of exposure. The final degradation product observed to be 2-nephthol was purified by High Pressure Liquid Chromatography (HPLC) and the molecular mass was computed by Gas Chromatography-Mass spectroscopy (GC-MS). The study revealed a cost effective and eco-friendly approach to degrade the toxic dyes into less toxic products by Staphylococcus sp. KU898286.