The aim of this work was to study chemical structures and biological activities of rhamnolipids produced by Pseudomonas aeruginosa B189 isolated from milk factory waste. The culture produced two biosurfactants, a and b, which showed strong activity and were identified as L-rhamnopyranosyl-(L)-rhamnopyranosyl-beta-hydroxydecanoyl-beta-hydroxyd ecanoate or Rha-Rha-C-10-C-10 and (L)-rhamnopyranosyl-(L)-rhamnopyranosyl-beta-hydroxydecanoyl-beta-hydrox ydodecanoate or Rha-Rha-C-10-C-12, respectively. Both compounds exhibited higher surfactant activities tested by the drop collapse test than several artificial surfactants such as SDS and Tween 80. Rhamnolipid a showed significant antiproliferative activity against human breast cancer cell line (MCF-7) at minimum inhibitory concentration (MIC) at 6.25 mu g/mL while rhamnolipid b showed MIC against insect cell line C6/36 at 50 mu g/mL. (c) 2006 Published by Elsevier Ltd.