On the early steps of cineol biosynthesis in Eucalyptus globulus

Samples of [4-H-2(1)]-1-deoxyxylulose (17a) and [2-C-13, 4-H-2(1)]-1-deoxyx lulose (17b), have been prepared by modification of known procedures and fe d in aqueous solution to twiglets of Eucalyptus globulus. The probes of cin eol (6) isolated from these experiments were analyzed by GC/MS, H-2- and C- 13-NMR techniques. In the experiments with lm, the formation of five isotop omers of 6 could be detected. Their structure and relative abundance demons trate that the C-13-label is incorporated to the same extent into the two C -5-units of 6, and that the H-2 label is retained to an extent of 57% in th e starter dimethylallyl-diphosphate unit (DMAPP; 12), but completely or alm ost completely lost in the unit derived from isopentenyl diphosphate (IPP; 11), in the elongation step which leads to geranyl diphosphate (GPP; 1). Th ese results confirm that the recently discovered mevalonate-independent pat hway to IPP and DMAPP is operative in the biosynthesis of cineol, and indic ate, together with previous finding, that, within this pathway, formation o f IPP and DMAPP occurs in independent rather than in sequential steps. In a ddition, the demonstration of different metabolic origins for the olefinic H-atoms of GPP (1), the aliphatic C-10-precursor of 6, paves the way for a realistic interpretation of the strikingly consistent but hitherto unexplai ned anomalies detected in the natural-abundance H-2-NMR spectra of (+)- and (-)-alpha-pinene and of (+)-limonene.