Pseudomonas aeruginosa was isolated from 20 years abandoned mine site of Itagunmodi Atakunmosa West, Ilesha, Nigeria. Atomic absorption Spectrophotometry (AAS) revealed Fe, Mn, Cr, Zn, Pb and Cr, while, Fe has the highest concentration range of 29-289 ppm in the analysed soil samples. Soil samples were enriched in R2b agar, serially diluted and pour plated. Four bacteria strains were isolated and identified using standard biochemical test. After routine biosurfactant screening by oil spreading and emulsification test, biosurfactant producing bacteria was confirmed as Pseudomonas aeruginosa. The partially purified biosurfactants were characterized with TLC and GC-MS analysis. The analyses indicated glycolipid biosurfactant specifically designated as Rhamnolipid-sa1 containing isopalmitic acid, hexadecanoic acid, methyl ester and hydroxylated fatty acid linked to decanoic acids. Iron removal potential of the extracted biosurfactant was studied and the result revealed that Rhamnolipid-sa1 effectively reduced iron (60.34%) and could be useful as alternative remediation tool for treatment of iron contaminated soil.