The new sulfated polyhydroxysteroid has been isolated from the Pacific starfish Mithrodia clavigera, collected from Maldives Islands and named as mitrotriol (I, Na-salt of (20S)-3beta,6alpha,20-trihydroxy-5alpha-cholest-9(11)-ene 3-O-sulfate). In addition six previously known compounds, including glycosides: echinasteroside B, granulatoside A, linckoside K, forbeside L and thornasterol sulfate A and cholesterol sulfate were isolated and identified. The structure of mitrotriol was elucidated by spectroscopic methods (mainly 2D NMR: 1H, 13C, DEPT, COSY-45, NOESY, HSQC and HMBC) and mass-spectrometry. For selected compounds, concentrations that showed cytotoxic activity against melanoma cells SK-MEL-28, SK-MEL-5 and RPMI-7951 were determined.