Glycosylceramide is a membrane lipid that has physiological functions in eukaryotic organisms. The presence of glucosylceramide has been confirmed in some yeast; however, the extent of the role of glucosylceramide in yeast is unknown. Thus, the extent of presence of glucosylceramide in yeast was surveyed using 90 strains of 24 genera. The strains were divided into two groups according to whether they had glucosylceramide (45 strains) or not (45 strains). The distribution of the ceramide glucosyltransferase gene (EC 2.4.1.80), which catalyzes glucosylation to a sphingoid lipid in glucosylceramide synthesis, and the phylogenetic classification of the strains were in agreement with those of glucosylceramide. Thus, the presence of glucosylceramide in yeast was caused by the presence of the gene involved in glucosylceramide synthesis and was closely associated with yeast evolution. Furthermore, the relationship between glucosylceramide presence and alkali tolerance of yeast was evaluated. The yeast with glucosylceramide tended to grow at higher pH, and a ceramide-glucosyltransferase-defective mutant from Kluyveromyces lactis did not grow at pH 8.5 even though the parent strain could grow under the same conditions. These results indicate that glucosylceramide in yeast might be a component that enables yeast to grow under alkali conditions.