The flotation performance of a copper ore sample in the presence of a rhamnolipid biosurfactant produced by Pseudomonas aeruginosa MA01 strain was studied. The rhamnolipid product was first investigated with respect to surface tension and frothing characteristics. Rhamnolipid showed a higher surface activity and frothability compared to DF-250 and MIBC frothers. Copper and molybdenum grades and recoveries were found to decrease as the biosurfactant concentration increased. In contrast, iron flotation was fairly improved in the presence of rhamnolipid. Results indicated that the use of rhamnolipid significantly increased the flotation rate of all minerals. As predicted by hydrophile-lipophile balance number/molecular weight diagram, rhamnolipid as a powerful surfactant increased the recovery of coarser particles to the concentrates. Although the use of rhamnolipids in copper ore flotation was not pleasing but the improved performance of iron minerals flotation may lead to its possible application in the dense mineral flotation such as lead and zinc. (C) 2011 Elsevier Ltd. All rights reserved.