Rhamnolipid biosurfactant enhancement of hexadecane biodegradation by Pseudomonas aeruginosa

Mutants of Pseudomonas aeruginosa that produce and do not produce rhamnolipid biosurfactant are used to investigate the influence of cell-associated biosurfactant on cellular association with the hydrocarbon-water interface and on hydrocarbon uptake. Rhamnolipid-nonproducing mutant 65E12 of P. aeruginosa is unable to grow in minimal media containing hexadecane as a carbon source in the absence of exogenously added surfactant. Mutant PG201::rhlR grows very slowly in the absence of exogenously added surfactants. Both mutants are deficient in the positive regulatory gene controlling the activation of rhamnolipid synthesis. 65E12 is a double mutant that is also deficient in lipopolysaccharide synthesis. However, growth on hexadecane may be restored to varying degrees when small amounts of purified rhamnolipids or the synthetic anionic surfactant alkyl benzene sulfonate (ABS) is added to the cultures. Rhamnolipid biosurfactant is shown to be approximately 9 times more effective than the structurally similar synthetic anionic surfactant ABS in solubilizing hydrocarbon into the aqueous phase. Physical characteristics of the rhamnolipid and ABS micelles as determined by laser light scattering are described to explain the greater effectiveness of the rhamnolipid in solubilizing hexadecane. The cellular attachment to hydrocarbon-water interfaces and cellular aggregation of the wild-type and mutant strains are examined in the presence and absence of rhamnolipid or synthetic ABS surfactants. Differences in observed hexadecane degradation rates are explained on the basis of emulsified hexadecane concentration, cell surface hydrophobicity, and cellular localization in the culture.