Steviamine, a new class of indolizidine alkaloid [(1R,2S,3R,5R,8aR)-3-hydroxymethyl-5-methyloctahydroindolizine-1,2-diol hydrobromide]

X-ray crystallographic analysis of the title hydrobromide salt, C10H20N+·Br−, of (1R,2S,3R,5R,8aR)-3-hydroxymethyl-5-methyloctahydroindolizine-1,2-diol defines the absolute and relative stereochemistry at the five chiral centres in steviamine, a new class of polyhydroxylated indolizidine alkaloid isolated from Stevia rebaudiana (Asteraceae) leaves. In the crystal structure, molecules are linked by intermolecular O—H⋯Br and N—H⋯Br hydrogen bonds, forming double chains around the twofold screw axes along the b-axis direction. Intramolecular O—H⋯O interactions occur.

Supplementary data and figures for this paper are available from the IUCr electronic archives (Reference: LH2918).  (Colegate et al., 1979), is a powerful inhibitor of α-mannosidases and has potential as a chemotherapeutic agent for the treatment of cancer (Lagana et al., 2006;Klein et al., 1999). l-Swainsonine 3, the enantiomer of 2, is a very powerful α-rhamnosidase inhibitor (Davis et al., 1996); 4 in which a methyl group is introduced into the piperidine ring is nearly 100 times more potent an inhibitor than 2 (Hakansson et al., 2008). Castanospermine 5, isolated from Castanospermum australe (Hohenschutz et al., 1981), is an inhibitor of some α-glucosidases and a potent inhibitor of dengue virus infection in vivo (Whitby et al., 2005); Celgosivir, a simple derivative of 5, is in development for the treatment of HCV infection (Durantel, 2009). Hyacinthacine A4 6, isolated from Scilla sibirica (Asano et al., 2000b;Yamashita et al., 2002), is the pyrrolizidine equivalent of steviamine 1. Many hyacinthacines have been isolated from a range of plants (Kato et al., 1999;Kato et al., 2007) and have attracted considerable attention from synthetic organic chemists (Sengoku et al., 2009;Donohoe et al., 2008). Steviamine 1 is unlikely to be the only naturally occurring indolizidine with a methyl branch which will provide similarly challenging synthetic targets.
As a natural product, the crystal was expected to be enantiopure and the Flack x parameter refined to 0.002 (10) (Flack, 1983). Analysis of the Bijvoet differences using within CRYSTALS (Betteridge et al., 2003) gives the Hooft y parameter as 0.023 (6), indicating that the probability that the configuration is incorrect allowing for the posibility of racemic twinning is less than 0.000001% (Hooft et al., 2008).
On examination of hydrogen bonding interactions in 1, the position of H51 initially seemed incorrect, lying between atoms O2 and O15. However, examination of the difference map indicates the presence of a peak believed to be a hydrogen atom which moves little on refinement suggesting the hydrogen bond is bifurcated (Fig. 2, Table 1). The molecules are linked together by three hydrogen bonds (two O-H···Br and one N-H···Br, Table 1) to form double chains around the twofold screw axes along the b direction (Fig. 3).

Experimental
Steviamine was isolated by a combination of strongly acidic cation, and strongly basic anion, exchange chromatography. The compound was retained on cation exchange resin (IR120) and was chromatographed on the anion exchange resin (CG400) from which it was eluted with water. Isolation was monitored using GC-MS of the trimethylsilyl-derivative (distinctive major ion at 314 amu). Steviamine was crystallized as its hydrobromide salt from ethanol. supplementary materials sup-2

Refinement
The H atoms were all located in a difference map, but those attached to carbon atoms were repositioned geometrically. The H atoms were initially refined separately with soft restraints on the bond lengths and angles to regularize their geometry (C-H in the range 0.93-0.98, O-H = 0.82 Å) and U iso (H) (in the range 1.2-1.5 times U eq of the parent atom), after which the positions were refined with riding constraints.
On comparison of F o and F c , it was apparent that for large values, of F o was noticably less than F c , so an extinction parameter was refined (Larson, 1970).      (4)