Traditional techniques for stereochemistry analysis have limitations; for example solution NMR spectroscopy has spatial limitation for the transference of stereochemical information, and suitable single crystals for X-ray analysis may not be available. Residual dipolar coupling (RDC) and powder X-ray diffraction (PXRD) are both techniques whose use is not yet widespread. We report a double-blind solution of the structure of jaborosalactol 24 (1), a new withanolide isolated from Jaborosa parviflora (Phil.) A. T. Hunziker et Barboza, by RDCs and PXRD, since conventional NMR spectroscopy experiments such as nuclear Overhauser effect (NOE) and 3 J analysis did not provide a unique solution.NOE, [1] frequently combined with 3 J coupling constants analysis, [2] is one of the most powerful tools for structural and conformational analysis by solution NMR spectroscopy. Unfortunately, the determination of relative stereochemistry between remotely located stereocenters becomes problematic arising from the limitations imposed by the 1/r 6 dependence of NOE (where r = internuclear distance). However, when molecules are forced to adopt a minor degree of alignment in solution and no longer tumble isotropically, a measurable fraction of the dipolar coupling (0.01-0.1 %) can be observed in the NMR spectrum. These so called residual dipolar couplings [3][4][5][6][7] (RDCs) contain important structural information of non-local character, since their values depend not only on the internuclear distances but also on the angles between the internuclear vectors and the external magnetic field. Hence, they provide information about the relative orientation between the internuclear vectors (e.g., H-H, H-C, H-N, C-C, etc.), regardless of the distance between them. Since the pioneering work by Courtieu and co-workers on the determination of relative configuration by RDCs using poly-gbenzyl-l-glutamate (PBLG)/CDCl 3 , [8,9] more alignment media compatible with organic solvents are now available. [10][11][12][13][14][15][16][17][18][19][20][21][22] In natural products in particular, the power of RDCs for the resolution of configurational problems has been tested against known molecules such as menthol, [23] cyclosporin, [15] ludartin, [22] strychnine, [21,24] sphaeropsidin A, [11] sagittamide A, [25] archazolid A, [26] sodium cholate, [27] and even used, in the case of the novel glycoside sucro-neo-lambertellin [28] for the determination of the unknown configuration in several stereocenters. Another case is the configuration of a synthetic a-methylene-g-butyrolactone that could not be resolved by conventional means. [29] At present, PXRD analysis can be used to determine the crystal structures of small organic molecules. [30] One fruitful technique is realspace simulated annealing, in which trial structures are generated and optimized using a set of structural parameters that define the location, orientation, and conformation of the molecules in the solid, consistent with bond lengths and angles from the known (or hypothesized) molecular stru...