Graduate Thesis Or Dissertation
 

The occurrence of abscisic acid in the dormant shoots of Douglas-fir [Pseudotsuga menziesii (Mirb.) Franco]

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  • Methanolic extracts of the dormant Douglas-fir shoots (buds, leaves, and stems) were fractionated by solvent partitioning (sodium bicarbonate-diethyl ether). The acidic portion of the extract was subjected to column (Sephadex LH-2O, Silica Gel and Polyvinylpyrrolidone), preparative thick-layer and gas-liquid chromatography. Fractions collected from chromatographic separations were collected and tested for biological activity by the standard straight-growth Avena bioassay. Three series of extracts, collected in 1969 (preliminary series), 1970-1971 (first series), and 1972 (second series) were analyzed. Results of the preliminary series (buds only) showed only the presence of growth inhibiting compounds. No growth promoters were observed. Analyses of the first series extracts (buds only) showed that the zones of maximum growth inhibition in column and preparative thick-layer chromatographic separations corresponded to the elution volumes and Rf values of abscisic acid. Preparative thick-layer and gas-liquid chromatography were used successfully to isolate abscisic acid from the extract. Confirmation of its presence was determined on the purified methylated (diazomethane) extract by mass spectrometry, conversion of abscisic acid into its C2-trans-isomer, and the positive results for specific color reaction. This is the first reported occurrence of abscisic acid in Douglas-fir. Abscisic acid was quantified in the remaining extracts of the first series. The occurrence of the C2-trans-isomer was not observed in any of the extracts. In the second series, abscisic acid in the dormant buds, leaves and stems was quantified. The results from this series showed high levels of abscisic acid in the buds (1.2 g/g fresh weight), leaves (0.347 g/g fresh weight), and stems (0.094 g/g fresh weight) for the fall collection (October 1972). All tissues showed the lowest levels of abscisic acid in early spring (February, March and April). Although the concentration of abs cisic acid was shown to follow the dormancy cycle in Douglas-fir, it appears to play a secondary role (if any)in the induction of dormancy in this species. The trend for abscisic acid concentration throughout the dormancy cycle appears to follow more closely the tree s capacity to biosynthe size the compound. One of the interfering compounds in the gas-liquid chromatographic analyses of abs cisic acid was determined by mass spectrometry and nuclear magnetic resonance to be dioctyl phthalate [di-(2-ethylhexyl)phthalate]. The natural occurrence of this phthalate ester in Douglas-fir is doubted and its presence was probably introduced as an artifact of the isolation procedure. The fact that it possessed inhibitory activity, however, stressed the importance for rigorous isolation procedures in abscisic acid analysis. Two other regions of growth inhibition were observed but no further chemical characterization was attempted.
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