Biosurfactants are of increasing interest due to their broad range of potential applications. A large variety of microbial surfactants is known at present, some of which may be used for specific applications. Towards the large scale industrial production of biosurfactants, the physiology, biochemistry and genetics of biosurfactant synthesis has to be well understood. A fully integrated process has to be developed, allowing high productivities under optimized conditions. In the past few years, we have investigated the molecular biology of rhamnolipid biosynthesis have been partially purified and characterized. The structural and regulatory genes encoding the rhamnolipid synthesis pathway have been isolated and characterized. The knowledge of the complex mechanisms involved in rhamnolipid synthesis facilitates the overproduction of these extracellular compounds. Furthermore, the transfer of the relevant genes into other species allows the production of rhamnolipids in heterologous hosts under controlled conditions. An integrated process for the production of rhamnolipids on an industrial scale has been developed. This process involves continuous cultivation under optimized media and growth conditions and makes use of refined methods of cell recycling, gas exchange and downstream processing, thus allowing high yields and productivities.