The biodegradation of hexadecane by five biosurfactant-producing bacterial strains (Pseudomonas aeruginosa UG2, Acinetobacter calcoaceticus RAG1, Rhodococcus erythropolis DSM 43066, R. erythropolis ATCC 19558, and strain BCG112) was determined in the presence and absence of exogenously added biosurfactants. The degradation of hexadecane by P. aeruginosa was stimulated only by the rhamnolipid biosurfactant produced by the same organism. This rhamnolipid did not stimulate the biodegradation of hexadecane by the four other strains to the same extent, nor was degradation of hexadecane by these strains stimulated by addition of their own biosurfactants. This suggests that P. aeruginosa has a mode of hexadecane uptake different from those of the other organisms. Rhamnolipid also enhanced the rate of epoxidation of the aliphatic hydrocarbon ␣,-tetradecadiene by a cell suspension of P. aeruginosa. Furthermore, the uptake of the hydrophobic probe 1-naphthylphenylamine by cells of P. aeruginosa was enhanced by rhamnolipid, as indicated by stopped-flow fluorescence experiments. Rhamnolipid did not stimulate the uptake rate of this probe in de-energized cells. These results indicate that an energy-dependent system is present in P. aeruginosa strain UG2 that mediates fast uptake of hydrophobic compounds in the presence of rhamnolipid.The bioavailability of sparingly soluble hydrophobic compounds for microbial conversion usually is very low and often limits their rate of degradation in aqueous and soil systems (2, 36). The use of biosurfactants has been found to enhance degradation in various cases. The degradation of long-chain alkanes, for example, by Pseudomonas aeruginosa can be stimulated by rhamnolipid, a biosurfactant produced by the organism (15, 37). Insight into the mechanism of rhamnolipid-enhanced degradation of hydrophobic compounds is desirable for the application of this organism or its biosurfactant in bioremediation or biotransformation.It has been reported that degradation of hexadecane by rhamnolipid-producing organisms is stimulated to a greater extent by rhamnolipid than by other biosurfactants (17) or synthetic surfactants (26). This is probably caused by the increase in cell surface hydrophobicity after extraction of lipopolysaccharides from the cellular envelope by rhamnolipid, which subsequently facilitates uptake via direct contact between cells and hydrocarbon droplets (1). In contrast, rhamnolipid failed to stimulate degradation of hydrophobic compounds by other strains or by mixed cultures which are not known to produce biosurfactants (15, 31). Thus, a high specificity seems to exist in the interaction between rhamnolipid and rhamnolipid-producing strains. It is not known whether such a specificity exists for other biosurfactant-producing bacteria and their biosurfactants in a similar way. Therefore, we have studied whether hexadecane biodegradation by P. aeruginosa and by four other biosurfactant-producing bacteria is stimulated by exogenous addition of their own and other biosurfactants. In orde...