Positive Identification of the Principal Component of a White Powder as Scopolamine by Quantitative One-Dimensional and Two-Dimensional NMR Techniques

An unidentified white powder collected as evidence in an intelligence investigation was characterized exclusively by nuclear magnetic resonance (NMR) analysis. A small fraction of the powder dissolved in D 2 O was subjected to a series of one- and two-dimensional techniques which were used to elucidate the molecular structure of the powder's major component and positively identify it as the scopolamine biotoxin. Quantitative one-dimensional experiments identified individual proton and carbon atom sites, and conventional 14 N spectroscopy detected a single nitrogen atom site. Heteronuclear single quantum coherence data correlated all protons to their directly bonded carbon atom, and together with the quantitative spectra, were used to determine the number of protons directly bonded to each carbon atom. The presence of a methyl, carboxyl, and a benzyl group was also identified from these data. Correlation spectroscopy detected a three proton and a nine proton J H,H network, representing a CH 2 CH moiety and seven carbon atom ring, respectively. These five elements were assembled into an almost complete molecular structure by using long-range, J-coupled, 'H- 13 C pairs detected by heteronuclear multiple bond correlation (HMBC) spectroscopy and 1 H- 1 H dipolar-coupled pairs found from nuclear Overhauser effect spectroscopy (NOESY) data. Additional oxygen atom sites were inferred from 1 H- 13 C correlation intensities in the HMBC spectra along with 'H and 13 C chemical shift values, or directly from NOESY correlations. Only a single oxygen atom site could not be inferred from NMR data, but its presence was inferred from comparisons to target analyte structures to complete the structure of the scopolamine molecule. To confirm these results, an ethanol/H 2 O solution of the powder was analyzed by direct infusion into an ion trap mass spectrometer. A prominent base signal was observed at m/z 304.1 amu, corresponding to the protonated molecular ion of scopolamine. Subsequently, the ion was selected and subjected to collision-induced dissociation, producing characteristic major MS/MS fragments at m/z 138.1 and 156.1. Comparisons of 1 H and 13 C chemical shift values and J H,H values measured from our NMR data were found to agree very favorably with previously reported values for scopolamine in D 2 O.