Specialized metabolism (secondary metabolism) in glandular trichomes of sweet basil (Ocimum basilicum L.) and accumulation of specialized metabolites (secondary metabolites) in rhizomes of turmeric (Curcuma longa L.) was investigated using proteomic and metabolomic approaches, respectively. In an effort to further clarify the regulation of metabolism in the glandular trichomes of sweet basil, we utilized a proteomics-based approach that applied MudPIT (multidimensional protein identification technology) and GeLC-MS/MS (gel enhanced LC-MS/MS) to protein samples from isolated trichomes of four different basil lines: MC, SW, SD, and EMX-1. Phosphorylation, ubiquitination and methylation of proteins in these samples were detected using X!tandem. Significant differences in distribution of the 755 non-redundant protein entries demonstrated that the proteomes of the glandular trichomes of the four basil lines were quite distinct. Correspondence between proteomic, EST, and metabolic profiling data demonstrated that both transcriptional regulation and post-transcriptional regulation contribute to the chemical diversity. One very interesting finding was that precursors for different classes of terpenoids, including mono- and sesquiterpenoids, appear to be almost exclusively supplied by the MEP (2-C-methyl-D-erythritol 4- phosphate) pathway, but not the mevolonate pathway, in basil glandular trichomes. Our results suggest that carbon flow can be readily redirected between the phenylpropanoid and terpenoid pathways in this specific cell type. To investigate the impact of genetic, developmental and environmental factors on the accumulation of phytochemicals in rhizomes of turmeric, we performed metabolomic analysis in a 2x2x4 full factorial design experiment using GC-MS, LC-MS, and LC-PDA. Our results showed that growth stage had the largest effect on levels of the three major curcuminoids. Co-regulated metabolite modules were detected, which provided valuable information for identification of phytochemicals and investigation of their biosynthesis. Based on LC-MS/MS data, 4 new diarylheptanoids were tentatively identified in turmeric rhizomes using Tandem-MSASC, a home-made software tool that automatically recognizes spectra of unknown compounds using three approaches. Based on our metabolomic results, we proposed two new strategies, “metabolomics-guided discovery” and “correlation bioassay”, to identify bioactive constituents from plant extracts based on information provided by metabolomic investigation.

Athrixia phylicoides (bush tea) is one of many plants from the Asteraceae family used as a traditional herbal medicine. With very few cultivated plants, natural growing plants currently serve as the main resource for plant material. The plant is not yet commercialised and its medicinal value is known and used only by a few people. With the long term aim at commercial scale propagation, this study consists of three parts. Firstly we developed a protocol for in vitro propagation of A. phylicoides. Secondly, the ultrastructure and morphology of leaves were studied microscopically and thirdly, comparisons were made between in vitro and ex vitro grown plants. Nodal segments of greenhouse plants were used to establish cultures. Better growth and less wilting was recorded on explants surface sterilised with NaOCl compared to Ca(OCl)2 after establishment. The addition of growth regulators IBA (indole-3-butyric acid) and BAP (6-benzylaminopurine) to the culture medium did not seem to affect the growth response of explants during the multiplication phase. Hyperhydricity was a problem throughout our trials. The development of hyperhydricity symptoms seems to be related to seasonal changes in the stock plant material used to initiate cultures, rather than the composition of growth medium or growth room temperatures. The occurrences of hyperhydricity symptoms were inconsistent and unpredictable. A rooting medium with added BAP and decreased sucrose levels resulted in a higher rooting percentage compared to the control medium, free from BAP and with a higher sucrose concentration, which yielded no rooting. However, in another experiment, in vitro rooting occurred spontaneously after subdividing and transfer of microshoots to fresh control medium. The addition of GA3 to the establishment medium (but not to the subsequently used multiplication and rooting media) yielded a slightly higher percentage of rooting. However, cultures initially established on GA3 medium yielded fewer roots per explant and roots were shorter than those of explants established on hormone free medium. The medicinal properties of plants are often linked to the production of essential oils. We hypothesised that the medicinal value of A. phylicoides can be linked to the production of the aromatic essential oils released by leaves. A microscopic study of leaves provided some preliminary insight of the mechanisms involved in the production of medicinally active products. Electron- and light microscopic examination of leaves were used to identify and study structures that are apparently involved in the production and secretion of essential oils. Two types of trichomes were identifyed – nonglandular and glandular trichomes. These glandular trichomes are multicellular with a subcuticular storage space and are present only on the adaxial surface of leaves. In the case of medicinal plants, it is essential that the medicinal properties of the plant are not altered by the method of propagation. This was our motivation for comparing the morphology and ultrastructure of leaves of plants that were grown in their natural environment to that of plants grown in vitro. Leaf surfaces of in vitro grown plants were smaller and the number of glandular trichomes per surface area was less on in vitro grown plants. There were no noticeable changes in the morphology of glandular trichomes. / Dissertation (MSc(Agric))--University of Pretoria, 2009. / Plant Production and Soil Science / unrestricted

Orientador: Silvia Rodrigues Machado / Resumo: O gênero Jacaranda Juss. (Bignoniaceae), um representante da tribo Jacarandeae, é constituído por 47 espécies, das quais 39 são endêmicas para o Brasil. Este gênero se destaca dos demais gêneros de Bignoniaceae principalmente por apresentar estaminódio alongado glandular e forte potencial etnobotânico e fitoquímico em várias espécies ocorrentes no Brasil. Estruturas secretoras são sítios de produção de substâncias bioativas, estudos sobre distribuição, morfologia e organização anatômica e sua composição química podem contribuir para conhecimento de Jacaranda no âmbito biológico, químico e farmacológico. Neste contexto, o objetivo deste estudo foi realizar uma descrição anatômica foliar de 24 espécies de Jacaranda ressaltando características capazes de agrupar espécies e comparar com ambientes de ocorrência e evidenciar características diagnósticas para as espécies. Dentre elas, também foi objetivo desse estudo, realizar uma prospecção das estruturas secretoras foliares e florais em seis espécies associando o morfotipo das glândulas com sua localização, organização anatômica e as principais classes de compostos produzidos. Para isso, usamos técnicas rotineiras para o processamento de material herborizado e material coletado fresco para estudos anatômicos, micromorfológicos de superfície, histoquímicos e analises de agrupamento. Características morfológicas e anatômicas foliares, tais como, o tamanho, forma, tipo da margem e textura dos foliólulos, assim como a estrutura da ala... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The genus Jacaranda Juss. (Bignoniaceae), a representative of the Jacarandeae tribe, consists of 47 species, 39 of which are endemic to Brazil. This genus stands out from the other genera of Bignoniaceae mainly for presenting elongated glandular staminode and strong ethnobotanical and phytochemical potential in several species occurring in Brazil. Secretory structures are sites of production of bioactive substances, studies on distribution, morphology and anatomical organization and their chemical composition can contribute to Jacaranda knowledge in the biological, chemical and pharmacological fields. In this context, the objective of this study was to perform a leaf anatomical description of 24 species of Jacaranda highlighting characteristics capable of grouping species and comparing with occurrence environments and highlighting diagnostic characteristics for the species. Among them, it was also the aim of this study, to prospect for leaf and floral secretory structures in six species associating the gland morphotype with its location, anatomical organization and the main classes of compounds produced. For this, we use routine techniques for the processing of herborized material and fresh collected material for anatomical, surface micromorphological, histochemical and cluster analysis studies. Leaf morphological and anatomical characteristics, such as the size, shape, type of margin and texture of the leaflets, as well as the structure of the wing and edge, as well as the t... (Complete abstract click electronic access below) / Doutor