Three new glycosidic antibiotics, glykenin (GK)-III A, B, and C (2a-c), were isolated as a mixture from a strain of Basidiomycetes sp. and identified as glycosides of C26-fatty acids and a diacetylated trisaccharide composed of glucose and two xyloses. The locations of the two acetyl groups were elucidated by secondary ion mass spectrometry (SIMS), double quantum filter (DQF), and relayed chemical shift correlation spectroscopy (COSY) spectral analysis.