Seeds of Cassia obtusifolia L. are known as homology of medicine and food material, which is a commonly consumed beverage in China. One new compound, 8-hydroxy-1,7-dimethoxy-3-methylanthracene-9,10-dione-2-O-β-d-glucoside (1), together with 11 known compounds, including seven anthraquinones (2-8), was isolated from the seeds. The 2D NMR data of compound 2 are reported for the first time. The structures of the compounds were established on the basis of 1D and 2D NMR, IR and HR-ESI-MS spectra. The cytotoxic activities of all the compounds against five cell lines (LO2, HCT-116, A549, HepG2 and SGC7901) were evaluated by using CCK8 methods. Compounds 1, 3 and 7 show moderate cytotoxicity towards HCT-116 cells compared with oxaliplatin.