A new xanthone glycoside, 8-hydroxy-3,5-dimethoxy-1-O-[α-L-arabinofuranosyl-(1 → 6)-β-D-glucopyranosyl]-xanthone (1) in addition to two known xanthones named 8-hydroxy-3,5-dimethoxy-1-O-β-Dglucopyranosyl-xanthone (2) and 1,8-dihydroxy-3,5-dimethoxyxanthone (methylbellidifolin) (3) were isolated from the methanol extract of aerial part of Centaurium spicatum, and tested for their cytotoxic activities against three different types of cell lines HeLa, THP-1and HL-60 cell lines. All compounds showed cytotoxicities to all cell lines, whereas showed a moderate activity against human monocytic cell line HL-60. Among three tested xanthones, the cytotoxic activity of compound 1 was stronger than those of other xanthones and the IC50 values for compound 1 were 3.22 (against THP-1), 8.67 (against HeLa) and 74.6 (against HL-60) μM, respectively.