e report thefirst complete structure elucida-tion of theent-kaurane diterpenoid glycoside atractyloside (1)by means of NMR and X-ray diffractometry techniques.Extensive one- and two-dimensional NMR experiments wereemployed to assign the proton and carbon signals of1, andcrystallography experiments established the configurations ofall stereogenic centers. Furthermore, we present a novelsemisynthetic route for the preparation of the highly cytotoxicaglycone derivative of1, 15-didehydroatractyligenin methylester (3). All compounds were tested for their antibioticactivity againstEnterococcus faecalis,Escherichia coli, and several strains ofStaphylococcus aureus, includingfluoroquinolone-resistant (SA1199B) and two epidemic MRSA (EMRSA-15 and -16) strains. Compound3exhibited moderate activity against allof theStaph. aureusstrains with an MIC value of 128 mg/L