The saponins, conjugated sterols, and free sterols of the sea cucumber Eupentacta fraudatrix were examined. A total of 85 steroids, twelve of them new, were identified in the free sterol, sulfated sterol, and sterol-xyloside fractions. The free sterol fraction contained 4α,14α-dimethylcholest-9(11)-en-3β-ol (6) and 14α-methylcholest-9(11)-en-3β-ol (7) together with 18 minor sterols. Examination of the aglycone moieties of the sterol-β-xyloside fraction afforded 31 different sterols. Cholestan-3β-ol (15) and 24-methylcholesta-7,22-dien-3β-ol (20) were the major sterols in this group. Cholestanol sulfate (74) and cholesterol sulfate (64) were identified as the major components among the 34 different sterol sulfates present. Finally, cucumariosides G 1 (1) , C 1 (2) , C 2 (3) , H (4) , and G 2 (5) were isolated from the saponin fraction. Radiolabeling experiments indicated that ther are two pathways of sterol biosynthesis in E. fraudratix. The first involves transformation of squalene to produce lanosta-9(11),24-dien-3β-ol (parkeol) which is subsequently demthylated to form 4α,14α-dimethylcholest-9(11)-en-3β-ol (6) and 14α-methylcholest-9(11)-en-3β-ol (7) . The second proceeds through squalene to lanosterol which is further metabolized to produce the triterpene saponins, 5α-cholest-7-en-3β-ol (19) and its xyloside (49) .