Quantitation and Localization of Bioactive Natural Products by Spectroscopic Methods

Abstract

This thesis describes spectroscopic analyses of plants which derive some, or all, of their commercial value from their secondary metabolites. The primary aim was to assess vibrational spectroscopy as a tool for rapid selection of commercially valuable crops. An equally important research direction was the exploration of the spatial distribution of plant secondary metabolites using Raman microscopy.

Raman spectra of powdered carrot (Daucus carota) were used to produce partial least squares regression models capable of quantitating carotenoids, but not polyacetylenes. The Raman and infrared spectra of powdered hops (Humulus lupulus) were used to quantitate α-acids, cohumulone, total bitter acids and xanthohumol. NIR spectra of the same samples could be used to predict the concentrations of α-acids and total bitter acids but not the concentrations of cohumulone or xanthohumol.

Raman microscopy was used to show that polyacetylene-rich oils exude from sub-dermal oil ducts in response to damage. This analysis was performed on fresh, sectioned carrot tissue in situ). The same technique was used to demonstrate that β-triketones and flavonoids are localized in the leaf oil glands of manuka (Leptospermum scoparium). Raman spectra of these oil glands could also be used to rapidly distinguish manuka chemotypes. Hops extracellular trichomes (lupulin) could also be used for rapidly chemotyping of hops cultivars and, more importantly, this analysis could be used to rapidly measure commercially important hops chemistry (i.e. xanthohumol concentrations and α:β acid ratios).

In the pursuit of these primary objectives, some related research avenues were encountered, and followed. These included: a comprehensive characterization of lupulin volatiles using solid-phase microextraction combined with gas chromatography, a similarly comprehensive analysis of lupulin bitter acids using proton nuclear magnetic resonance spectroscopy; isolation and characterization of three new β-triketones and a new flavanone from manuka; and the application of chemometrics to chromatographic and nuclear magnetic resonance data-sets.