Rhamnolipids find broad applications as natural surfactants, emulsifiers, and antibiotics because of their low toxicity, high biodegradability and environmental soundness. In this study, a pH stage-controlled process of fermentation of rhamnolipids by Pseudomonas aeruginosa O-2-2 was established. A yield of 24.06 g/L in batch fermentation was achieved in a 5-L fermentor via the optimization of stirring speed. By controlling pH, rhamnolipid production was increased to 28.8 g/L, an improvement of 19.7%, and more substrate was converted to rhamnolipids rather than to biomass. Fermentation kinetics models for cell growth, product synthesis and substrate consumption based on the pH stage-controlled fermentation indicated that rhamnolipid production could be further improved by fed-batch fermentation. Rhamnolipid production reached 70.56 g/L, an improvement of 193%, in the pH stage-controlled fed-batch fermentation when the stirring speeds was controlled at 500 rpm and the fermentation temperature was maintained at 30 degrees C. (C) 2012 Elsevier Ltd. All rights reserved.