The influence of configuration at C-20 on rotation about the 17(20)-bond in steroids and euphoids was examined by x-ray crystallographic studies of the C-20 epimers euphol and tirucallol. The H atom on C-20 was in back next to C-18 in the crystal structures of both of the compounds, and C-22 was found to be cis-oriented ("left-handed") to C-13 in euphol and trans-oriented to it ("right-handed") in tirucallol.The results, which are consistent with the known left-handed crystal structure of 24(25)-dihydroeuphol and right-handed crystal structure of cholesterol and other natural sterols, lend further credence to the earlier suggestion that rotational isomerism at the 17(20)-bond can arise in C-20 epinters and that there is preference for an arrangement with the 20-H atom adjacent to C-18.It is known from x-ray data (1-3) that in the crystalline state, sterols with the natural configuration (20a-H atom, usually 20R in the sequence rule) have a conformation about the 17(20)-bond such that, in the usual view of the molecule, C-22 is to the right. As we suggested earlier (4, 5), the preference for this "right-handed" rotational isomer probably derives from its having the smallest of the groups on C-20 (the H atom) in front and, therefore, adjacent to C-18 in a pseudo-1,3-diaxial fashion. If this were the correct explanation, the H atom on C-20 should remain adjacent to C-18 after inversion. This would place C-22 on the left ("left-handed" isomer)We undertook a study of this stereochemistry, because the ability of the sterol side chain to assume a right-handed conformation without having a methyl group in front seems to be of biological importance. There is a clear correlation between the biological activities of (20R)-sterols (which can have this stereochemistry) and (E)-17(20)-dehydrocholesterol with C-22 fixed to the right (5)(6)(7)(8)(9)(10)(11). Similarly, the activities of (20S)-sterols correlate with (Z)-17(20)-dehydrocholesterol (5-10). If only one of the 17(20)-dehydrosterols is active, it is the right-handed (E)-isomer, and then of the C-20 epimers only the one with the (20R)-configuration is active (5-10). These correlations have been found in the ability of sterols to perform what is thought to be the bulk membrane function in yeast (5, 6), to be metabolized by a protozoan (7,8), and to induce formation of oospores in an oomycete (9, 10). If, on the other hand, both of the 17(20)-dehydrosterols are active, as they are in depressing hepatic cholesterol synthesis after being fed to mice (11), then both of the C-20 epimers of cholesterol are active (11).To determine whether inversion of the C-20 configuration of sterols is actually accompanied in the solid state by transfer of C-22 from the right to the left side of the molecule, we have tried to grow large enough crystals of 20-epicholesterol, prepared as described (12), to permit x-ray analysis.Unfortunately, crystals of adequate size have not been obtained, so we turned our attention to the naturally occurring (13) C-20 epirners euphol and tirucallol...