Aims: To isolate and characterize biosurfactants produced by a thermotolerant yeast isolated in Thailand. Materials and Results: Yeast strains isolated from plant material in Thailand were first screened for the ability to produce lipase and biosurfactant. A strain Y12, identified as Candida ishiwadae by physiological tests, survived at 45degreesC and produced relatively large amounts of biosurfactants. From the culture filtrate of this strain, two glycolipid biosurfactants, a and b, were purified by solvent fractionation, silica gel and ODS column chromatographies. Compounds a and b were determined to be monoacylglycerols; 1-linoleylglycerol and 1-oleylglycerol, respectively. Both compounds exhibited higher surfactant activities tested by the drop collapse test than several artificial surfactants such as sodium dodecyl sulphate. Conclusions: Glycolipid biosurfactants produced by a thermotolerant yeast, C. ishiwadae were characterized to be monoacylglycerols which exhibited high surfactant activities. Significance and Impact of the Study: A thermotolerant yeast strain, C. ishiwadae, could be a potential candidate for producing monoacylglycerols which are useful in industrial applications.