Chromatographic analyses of crude leaf extracts of Flaveria linearis L. revealed the presence of four acetylenic monothiophenes. Three of these metabolites were purified and structurally characterized by UV, NMR, IR, and GC-MS. Germination, growth, and survival/mortality studies with and without UVA, the activating wavelengths of these metabolites, were conducted with the crude leaf extracts and the purified compounds (taken individually and combined) against selected crop species (lettuce, radish, and carrot). Results suggest that acetylenic metabolites are phytotoxic against lettuce, carrot, and radish, but with variability in response among species. These variations in sensitivity and the allelopathic potential of F. linearis is discussed.

Submitted in fulfillment for the Degree of Doctor of Technology: Biotechnology, Durban University of Technology, 2010. / Ceratatheca triloba (Bernh.) E. Mey. Ex Hook. f. is one of four species that is common to the summer rainfall areas in South Africa, especially the grasslands. It is used in traditional medicine to treat stomach cramps, nausea, fever and diarrhea. Like many other plants used in the traditional medicine system, these uses are not justified through scientific investigations. This study was undertaken to characterize the functionality of the main bioactive compounds from Ceratatheca triloba. This was achieved by isolating and identifying predominant chemicals from the non polar extracts using conventional chromatography techniques. Once identified the crude extracts and identified compounds were tested for their antimicrobial, anti-oxidant activity, anti-inflammatory activity and anticancer activity. This was followed by investigating the safety of the crude extracts and the purified compounds by the Brine shrimp lethality assay, and its toxicity to HepG2 cells and the Salmonella mutagenecity test. For large scale production, we set up a protocol to produce 9, 10 anthracenedione in a cell suspension culture system. Following the complete chemical profile of the roots, stems, flowers and leaves the predominant compounds were isolated, characterized and identified by UV-Vis, IR, EI-LCMS and NMR (COSY, HMQC, HMBC and DEPT). Three anthraquinone derivatives and one steroid, 9, 10 anthracenedione, 1-hydroxy-4-methylanthraquinone, 5, 8-dimethoxy-2, 3, 10, 10a-tetrahydro-1H-phenanthrene-4, 9-dione and androst-5-ene-3, 17, 19-triol were determine by analysis of spectral data (UV, 1H NMR, 13C NMR and EI-LC-MS) 9, 10 anthracenedione and 1 hydroxy-4-methylanthraquinone showed antibacterial activity against S.aureus, M. luteus, B cureus and E. coli. Due to the synergistic effect of the individual compounds, the crude extract exhibited good potency (>500) against S.aureus and M. luteus, medium potency against E. coli. and S. typhimurium (<100) and very low potency against B cureus (<10). Although a similar trend was observed for 9, 10 anthracenedione and 1 hydroxy-4-methylanthraquinone unlike the crude extract. A very low potency against S.aureus for 9, 10 anthracenedione and a high potency for 1 hydroxy-4-methylanthraquinone. Thus 9, 10 anthracenedione is an effective drug against E. coli and S. typhimurium and 1 hydroxy-4-methylanthraquinone is effective against S.aureus and M. luteus. The crude root extracts and 9, 10 anthracenedione, 1 hydroxy-4-methylanthraquinone, 8-dimethoxy-2, 3, 10, 10a-tetrahydro-1H-phenanthrene-4 showed a ± 50% reduction of the free radicals. No anti-inflammatory activity was observed. The purified extracts showed moderate toxicity against HepG2 cells at high concentrations and no toxicity was observed against brine shimp larvae. No mutagenecity was observed with the crude extracts using the Ames test. All purified and crude extracts showed potent inhibition of the human topoisomerase II enzyme. In conclusion, although this study does not indicate any relationship to its traditional usage it provides valuable information that paves a way for commercial exploitation of C. triloba. 9, 10 anthracenedione and 1 hydroxy-4-methylanthraquinone can be used as antibacterial agents. Their antioxidative potential can be exploited for anti-cancer as in many cancers reactive oxygen species are implicated in the aetiology of these cancers. Furthermore, in this study 9, 10 anthracenedione was produced from both callus cultures and cell suspension cultures. This compound demonstrates potent anti-topoisomerase II activity which is vital to cancer treatment. Thus, the synergistic effect of 9, 10 anthracenedione and 1 hydroxy-4-methylanthraquinone as antibacterial, anti-oxidative and anti-cancer compounds demonstrate the importance of C. triloba. / Centre for Research Capacity Development ; National Research Foundation

Submitted in complete fulfillment for the Degree of Master of Technology: Biotechnology, Durban University of Technology, 2012. / Many secondary metabolites that have been extracted from medicinal plants have been used as source of clinical drugs. However, the concentration of the active metabolites in plants is generally low. An attractive alternative for producing these important secondary metabolites is via plant tissue culture technology. More particularly, the genetic transformation of a plant tissue by Agrobaterium rhizogenes has been employed for producing high yields of secondary metabolites. In a previous study, three structurally similar anthraquinones: 9,10-Anthracenedione, 1-Hydroxy-4-methylanthraquinone and 5,8-Dimethoxy-2,3,10,10a-tetrahydro-1H,4aH-phenanthrene-4,9-dione, and one steroid; Androst-5-ene-3, 17, 19-triol were isolated from the root extracts of C. triloba. The anthraquinones have shown to exhibit the anticancer mechanism which involves the inhibition of the activity of the human topoisomerase II enzyme that transforms supercoiled DNA to linear DNA. However, these anthraquinones were found in very low concentrations. Therefore, in this study we used plant cell and tissue culture systems (cell suspension, shoot and hairy root cultures) of C. triloba to increase the production of anthraquinones. Since the establishment of C. triloba in vitro plant systems required a source sterile explants, a protocol that involved the use of NaCIO was optimized for the sterilization and subsequent germination of C. triloba seeds which were micro-propagated into shoot cultures. These cultures provided a source explants for the induction of callus and hairy root cultures. The biomass of these plant cell and tissue cultures were subsequently bulked up for the extraction for anthraquinones and the yields were compared followed by fractionation and identification of the major compounds. The bioactivity of the fractions was evaluated by testing their cytotoxicity on cancer cells and anti-topoisomerase activity. The sterilization protocol that provided sterile seeds was found to be a solution of 30% NaCIO at an exposure time of 10 minutes. From the sterilized seeds shoot cultures were established on MS medium. The leaf explants of the shoot cultures were then used to induce callus cultures which subsequently were transferred to liquid medium whereby the total biomass of suspension cultures increased from 4 g to 134.18 g (wet weight). Also hairy roots cultures were established from stem explants with a low cell density inoculum of A. rhizogenes at a transformation efficiency of 73%. The growth of these hairy roots was slow in hormone free medium. This was overcomed with the use NAA and IAA which increased the xvii biomass from 1.03 g in the control culture (without hormone) to 23.91 g and 46.13 g respectively. An evaluation of the anthraquinones in the field root and hairy root, cell suspension and shoot culture extracts was carried out by using their Thin Layer Chromatography profiles and the High Performance Liquid Chromatography profiles as well as the standards, 9,10-Anthracenedione and 1-Hydroxy-4-methylanthaquinone. TLC analysis showed that the RF values of the fractions CT01 and CT02 matched the RF values of anthraquinones standards while HPLC analysis revealed that hairy root cultures supplemented with IAA (125.03 μg.mg-1) or NAA (98.25 μg. mg-1) produced a higher concentration of anthraquinones than the control culture (without hormone) (13.33 μg.mg-1), the field roots (33.51 μg. mg-1) and the shoot (3.23 μg.mg-1) and cell suspension cultures (13.17 μg.mg-1). Due to co-elution of the compounds in HPLC analysis, six fractions were isolated by Preparative Thin Layer Chromatography from the hairy root extract (obtained from the culture supplemented with NAA) and were coded as CT01, CT02, CT03, CT04, CT05 and CT06. The compounds in these fractions were identified by Electron Ionization-Liquid chromatography-Mass Spectroscopy and it was found that the hairy roots produced one acridone derivative; 5-Methoxy-2-nitro-10H-acridin-9-one, one naphthoquinone derivative; 2H-Naphto[2,3-b]pyran-5,10-dione,3,4-dihydro-2,2-dimethyl- and seven anthracenedione derivatives. These were: i) 5,8-Dimethoxy-2,3,10,10a-tetrahydro-1H,4aH-phenanthrene-4,9-dione, ii) 9,10-Anthracenedione, 2-methyl-, iii) 1-Hydroxy-4-methylanthraquinone, iv) 9,10-Anthracenedione, 2-ethyl-, v) 1,5-Diaminoanthraquinone, vi) Phenanthrene, 3,6-dimethoxy-9-methyl-, vii) 9,10-Anthracenedione, 1,4-dimethyl-. Fractions CT01 (5,8-Dimethoxy-2,3,10,10a-tetrahydro-1H,4aH-phenanthrene-4,9-dione, 9,10-Anthracenedione, 2-methyl- and 1-Hydroxy-4-methylanthraquinone) and CT02 (9,10- Anthracenedione, 2-ethyl-) were cytotoxic to the DU-145 cancer cell line at concentrations of 125 μg.mg-1 to 1000 μg.mg-1. These fractions also showed anti-topoisomerase activity as they inhibited the conversion of supercoiled DNA into linear DNA. In conclusion this is the first study that describes the transformation of C. triloba by A. rhizogenes mediated transformation and compares the production of anthraquinones in C. triloba hairy roots to the field roots, shoot and cell suspension cultures. This study has xviii indicated that hairy root cultures is a high-yielding production system for anthraquinones (5,8-Dimethoxy-2,3,10,10a-tetrahydro-1H,4aH-phenanthrene-4,9-dione, 1-Hydroxy-4-methylanthraquinone, 9,10-Anthracenedione, 2-methyl- and 9,10- Anthracenedione, 2-ethyl-) which could have the potential to be used in cancer therapy. In addition the discovery of C. triloba hairy roots having the biosynthetic capacity to synthesize five valuable anthraquinone derivatives that are not found the field roots has also been revealed. / National Research Foundation.

Nejedly, J.: Nutraceuticals 5. Compounds usable as an adjuvant in the treatment of Alzheimer's disease. Diploma thesis, Charles University in Prague, Faculty of Pharmacy in Hradec Kralove, Department of Pharmaceutical Botany and Ecology, Hradec Kralove 2014, 102 p. This literature review, dealing with the pathophysiology of Alzheimer's disease and possible use of nutraceutical products as an adjuvant in the therapy of the disease, has been done by analysis of foreign and domestic literature. In the chapter Alzheimer's disease, various pathological processes, leading to the development of the disease and also risk factors, clinical picture and a brief indications of currently used treatment, were introduced. The main part of thesis called Nutraceuticals has introduced a comprehensive overview and categorization of compounds (plant extracts) which can be used in influencing of Alzheimer's disease by the mechanism of action. In the chapter Nutraceuticals in Czech Republic, various product and suplements were listed, which are available on the market in Czech Republic and could be used as an adjuvant in the therapy of Alzheimer's disease. Finally, it was shown the evaluation (pros, cons, potential interaction) of substances and supplements, which have been state in the thesis, with a proposal for their...

<p>Over the past 200 years, the Earth's atmospheric carbon dioxide (CO₂) concentration has increased by more than 35%, and climate experts predict that CO₂ levels may double by the end of this century. Understanding the mechanisms of resource management in plants is fundamental for predicting how plants will respond to the increase in atmospheric CO₂. Plant productivity sustains life on Earth and is a principal component of the planet's system that regulates atmospheric CO₂ concentration. As such, one of the central goals of plant science is to understand the regulatory mechanisms of plant growth in a changing environment. Short-lived positron-emitting radiotracer techniques provide time-dependent data that are critical for developing models of metabolite transport and resource distribution in plants and their microenvironments. To better understand the effects of environmental changes on resource transport and allocation in plants, we have developed a system for real-time measurements of metabolite transport in plants using short-lived positron-emitting radiotracers. This thesis project includes the design, construction, and demonstration of the capabilities of this system for performing real-time measurements of metabolite transport in plants.</p><p>The short-lived radiotracer system described in this dissertation takes advantage of the combined capabilities and close proximity of two research facilities at Duke University: the Triangle Universities Nuclear Laboratory (TUNL) and the Duke University Phytotron, which are separated by approximately 100 meters. The short-lived positron-emitting radioisotopes are generated using the 10-MV tandem Van de Graaff accelerator located in the main TUNL building, which provides the capability of producing short-lived positron-emitting isotopes such as carbon-11 (¹¹C; 20 minute half-life), nitrogen-13 (¹³N; 10 minute half-life), fluorine-18 (¹⁸F; 110 minute half-life), and oxygen-15 (¹⁵O; 2 minute half-life). The radioisotopes may be introduced to plants as biologically active molecules such as ¹¹CO₂, ¹³NO₃^-, ¹⁸F^--[H₂O], and H₂<sup>15<\sup>O. Plants for these studies are grown in controlled-environment chambers at the Phytotron. The chambers offer an array of control for temperature, humidity, atmospheric CO₂ concentration, and light intensity. Additionally, the Phytotron houses one large reach-in growth chamber that is dedicated to this project for radioisotope labeling measurements.</p><p>There are several important properties of short-lived positron-emitting radiotracers that make them well suited for use in investigating metabolite transport in plants. First, because the molecular mass of a radioisotope-tagged compound is only minutely different from the corresponding stable compound, radiotracer substances should be metabolized and transported in plants the same as their non-radioactive counterparts. Second, because the relatively high energy gamma rays emitted from electron-positron annihilation are attenuated very little by plant tissue, the real-time distribution of a radiotracer can be measured <em>in vivo</em> in plants. Finally, the short radioactive half-lives of these isotopes allow for repeat measurements on the same plant in a short period of time. For example, in studies of short-term environmental changes on plant metabolite dynamics, a single plant can be labeled multiple times to measure its responses to different environmental conditions. Also, different short-lived radiotracers can be applied to the same plant over a short period of time to investigate the transport and allocation of various metabolites.</p><p>This newly developed system provides the capabilities for production of ¹¹CO₂ at TUNL, transfer of the ¹¹CO₂ gas from the target area at TUNL to a radiation-shielded cryogenic trap at the Phytotron, labeling of photoassimilates with ¹¹C, and <em>in vivo</em> gamma-ray detection for real-time measurements of the radiotracer distribution in small plants. The experimental techniques and instrumentation that enabled the quantitative biological studies reported in this thesis were developed through a series of experiments made at TUNL and the Phytotron. Collimated single detectors and coincidence counting techniques were used to monitor the radiotracer distribution on a coarse spatial scale. Additionally, a prototype Versatile Imager for Positron Emitting Radiotracers (VIPER) was built to provide the capability of measuring radiotracer distributions in plants with high spatial resolution (~2.5 mm). This device enables detailed quantification of real-time metabolite dynamics on fine spatial scales.</p><p>The full capabilities of this radiotracer system were utilized in an investigation of the effects of elevated atmospheric CO₂ concentration and root nutrient availability on the transport and allocation of recently fixed carbon, including that released from the roots via exudation or respiration, in two grass species. The ¹¹CO₂ gas was introduced to a leaf on the plants grown at either ambient or elevated atmospheric CO₂. Two sequential measurements were performed per day on each plant: a control nutrient solution labeling immediately followed by labeling with a 10-fold increase or decrease in nutrient concentration. The real-time distribution of ¹¹C-labeled photoassimilate was measured <em>in vivo</em> throughout the plant and root environment. This measurement resulted in the <em>first</em> observation of a rapid plant response to short-term changes in nutrient availability via correlated changes in the photoassimilate allocation to root exudates. Our data indicated that root exudation was consistently enhanced at lower nutrient concentrations. Also, we found that elevated atmospheric CO₂ increased the velocity of photoassimilate transport throughout the plant, enhanced root exudation in an annual crop grass, and reduced root exudation in a perennial native grass.</p> / Dissertation

Thesis (Ph. D.)--Duke University, 2008.

Thesis (Ph.D.) - University of KwaZulu-Natal, Pietermaritzburg, 2009. / Full text also available online. Scroll down for electronic link.

Thesis (M.Phil.) - University of Queensland, 2006. / Includes bibliography.

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany and Ecology Candidate: Jana Puzyrevská Supervisor: PharmDr. Anna Hošťálková, Ph.D. Title of Thesis: Biological activity of plant metabolites XXXIV. Alkaloids from the herb of Glaucium flavum CRANTZ and their impact on human cholinesterases. Key words: Glaucium flavum Crantz, cataline, N-methyllaurotetanine, norchelidonine, protopine, AChE, BuChE. Alzheimer's disease, the most widespread neurodegenerative disease, causes decrease of cognitive functions and dementia. The most effective therapeutic approach is the application of central cholinesterase inhibitors, which alleviate cholinergic deficit in brain and thus improve memory. Currently, intensive investigation of new active compounds including natural substances is carried on. Within the preliminary testing, alkaloid extract from Glaucium flavum Crantz herb showed promising inhibition of human cholinesterases, so it was selected for further examination. The primary alkaloid extract was acquired from dried flowering herb by extraction with ethanol and subsequent liquid extraction at different pH. This extract was treated by preparative thin layer chromatography. The structure of alkaloids was determined by spectrometric methods (MS, NMR) and their optical...

D.Phil. (Biochemistry) / Plants are able to recognise and respond to signals from the environment through a complex array of biochemical pathways, which enables them to deter pathogenic micro-organisms and herbivores. Thousands of different structures of low-molecular weight organic compounds / natural products can be produced through an inducible chemical defence system; that can be manipulated for biotechnological purposes. The importance of natural products in medicine, agriculture and industry has led to numerous studies such as this, to understand the biosynthesis and biological activity of these substances...