Production of Glycolipid Biosurfactants, Mannosylerythritol Lipids, by Pseudozyma siamensis CBS 9960 and Their Interfacial Properties

The search for a novel producer of glycolipid biosurfactants, mannosylerythritol lipids (MELs), was undertaken on the basis of the analysis of ribosomal DNA sequences of yeast strains of the genus Pseudozyma . In the course of the investigation, Pseudozyma siamensis CBS 9960, which is closely related to Pseudozyma shanxiensis , a known MEL-C producer but with a different morphology, was found to accumulate a large amount of glycolipids. On thin layer chromatography, the extracellular glycolipids showed nearly the same spots as those of the MELs produced by P. shanxiensis . However, the result of high-performance liquid chromatography analysis revealed that the present strain has a much higher glycolipid production yield than P. shanxiensis. From the structural characterization by 1 H and 13 C NMR, the major glycolipid (more than 84% of the total) was identified as a mixture of 4- O -[(2′,4′-di- O -acetyl-3′- O -alka(e)noyl)-β- d -mannopyranosyl]- d -erythritol and 4- O -[(4′- O -acetyl-3′- O -alka(e)noyl-2′- O -butanoyl)-β- d -mannopyranosyl]- d -erythritol, both of which are types of MEL-C. The present MEL-C possessed a short-chain acid (C 2 or C 4 ) at the C-2′ position and a long-chain acid (C 16 ) at the C-3′ position of the mannose moiety, and thus, the hydrophobic part was considerably different from that of conventional MELs, which mainly possess two medium-chain acids (C 10 ) at the C-2′ and C-3′ positions. Under optimal growth conditions with safflower oil in a shake culture, the total amount of MELs reached approximately 19 g/ l after 9 d at 25°C. We further investigated the interfacial properties of the present MEL-C, considering its unique hydrophobic structure. The observed critical micelle concentration (CMC) and the surface tension at the CMC of the MEL were 4.5×10 −6 M and 30.7 mN/m, respectively. In addition, on a water penetration scan, the MEL efficiently formed the liquid crystal phases such as hexagonal (H) and lamella (L a ) at a wide range of concentrations. These results demonstrated that the newly identified MEL-C produced by P. siamensis exhibits not only high surface activity but also excellent self-assembling properties, and should facilitate the development of promising yeast biosurfactants.