Charles University, Faculty of Pharmacy in Hradec Kralové Department of Social and Clinical Pharmacy Author Anna Faltysová Supervisor PharmDr. Jitka Pokladníková, Ph.D. Consultant PharmDr. Julie Zubrová Title of Diploma Thesis Analysis of the risks associated with using dietary supplements by patients in pre-operative period Objectives: The thesis deals with the problem of the dietary supplement use by patients in the time before surgery. The aim of the thesis is to evaluate the prevalence of dietary suplement use by patients before surgery. Another aim is to point to potential danger which could endanger the patient while irrational using of the dietary supplement before the medical interventions (interactions of dietary supplement, side effects of dietary supplement). Methodology: The questionnaire research was conducted in order to obtain the data. It was counducted between 9th May 2017 till 30th June 2018 at 9 departments of the University Hospital in Hradec Králové. In total 332 patients in the time before surgery were asked and 257 completed questionnaires were analysed. Results: From the results of the study we can see the prevalence of the dietary supplement use by the patients in the time before surgery, which amounts to 42 %. It was stated that most of the patients does not inform their...

Thesis (M.Sc. (Biochemistry)) --University of Limpopo, 2011 / Dicerocaryum senecioides is a crawling herb that is found growing mostly in sandy areas of southern and south-eastern Africa and its small, hairy leaves have been used over the years as food, shampoo, and for treatment of various ailments. In this study, the dichloromethane (D2) fraction was prepared from a crude methanol extract of D. senecoides leaves, and its effect on the proliferation of RAW 264.7 murine macrophages was investigated. Treatment of the macrophages with the extract resulted in a dose- and time-dependent decrease in cell viability as determined by the MTT assay and real time cell analysis. Cytotoxicity of the D2 fraction on the macrophages was demonstrated to be due to apoptosis by staining the cells with DAPI nucleic acid stain. Anti-inflammatory activity of D2 fraction on RAW cells was determined by evaluating intracellular ROS production by the DCFH-DA fluorescent assay. Cells treated with the D2 fraction and stimulated with PMA were found to have a lower fluorescence intensity compared to untreated, stimulated cells; thus mimicking the response observed in the resting cells. The percentage fluorescence in untreated, stimulated cells doubled, while no significant change was observed in the D2-treated cells. The effect of the D2 fraction on iNOS activity was also assessed. The fraction reduced the NO synthesised by iNOS in cells treated with the D2 fraction and stimulated with LPS dose-dependently. The D2 fraction was further fractionated by semi-preparative HPLC; and thin layer chromatography was used to analyse phytocompounds of the 96 HPLC sub-fractions as well as to screen these sub-fractions for anti-oxidative activity. Sub-fractions 1-7 and 33-39 showed an intensely pronounced DPPH-scavenging compound and this scavenging ability was confirmed by a quantitative DPPH assay that provided parallel results. The reducing potential of the sub-fractions was assessed by evaluating their Fe3+-reducing ability through the FRAP assay. Sub-fractions 1-7 and 33-39 displayed remarkable reducing potential. Taken together with the DPPH-scavenging activity, these findings suggest that HPLC sub-fractions 1-7 and 33-39 possess a compound(s) with impressive antioxidant activity. These findings merit the D2 fraction as an extract that can be used to control chronic inflammation as it does not only inhibit free radical production, but also scavenges excessive ROS and has the ability to induce apoptosis in the macrophages responsible for dysregulated production of the free radicals. The extract also has commendable chemoprotective and chemotherapeutic potential as it demonstrated pro-apoptotic activity along with prevention of excess free-radical production. / National Research Foundation and the University of Limpopo Research Office

Thesis (Ph.D. (Biochemistry)) --University of Limpopo, 2009 / Commelina benghalensis Linn is used in traditional medicine in several Asian and African countries for the treatment of various ailments such as stomach irritations, burns, sore throat and feet, diarrhoea and as an anti-inflammatory agent. Recently, our laboratory showed that the crude methanolic extract of Commelina benghalensis L (CMECB) exhibits growth inhibitory and proapoptotic effects in Jurkat T and Wil-2 NS cancer cell lines. In this study, the precise molecular mechanism(s) associated with CMECB-induced growth inhibitory and apoptosis inducing effects in Jurkat T and Wil-2 NS cell lines were investigated. This was achieved by investigating the effects of the extract on the cell division cycle distribution profile as well as its effects on various cell division cycle and apoptosis regulatory genes. Ground stems of C. benghalensis L were extracted with absolute methanol to obtain a crude extract. To assess the effect of CMECB on cancer cell growth, experimental cell cultures were exposed to various concentrations (0 to 600 μg/ml) of CMECB for up to 72 hours. The results demonstrated a significant reduction in cell viability and inhibition of proliferation of experimental cell cultures as determined by the trypan blue dye exclusion assay and the Coulter counter method, respectively. Analysis of nuclear morphological changes in cells stained with Hoechst 33258 confirmed apoptosis as the mode of cell death that is associated with the growth inhibitory effects of CMECB in both the Jurkat T and Wil-2 NS cell lines. This assertion was based on the observed presence of nuclear morphological changes such as chromatin condensation and fragmentation and apoptotic bodies in cells exposed to CMECB. In order to get an insight on the pro-apoptotic mechanisms of CMECB, Western blot xxi and quantitative real-time PCR (qrt-PCR) were used to investigate the expression profiles of various apoptosis and cell division cycle regulatory genes. Qrt-PCR results showed a lack of a clear up- and/or down-regulatory effects of CMECB on the mRNA expression levels of bax and bcl-2 in both Jurkat T and Wil-2 NS cells. Western blot analyses demonstrated that CMECB induced apoptosis by facilitating Bax protein translocation from the cytosol to the mitochondria in both Jurkat T and Wil-2 NS cells. In addition, CMECB down-regulated Bcl-2 protein expression which, as a result, led to the shift in the Bax/Bcl-2 protein ratio at certain time points and concentration in both Jurkat T and Wil-2 NS cells. The modulation of the Bcl-2 family members led to mitochondrial cytochrome c release into the cytosol and activation of caspases-9 and -3; this was also confirmed by caspase activity assays and eventual degradation of PARP. Furthermore, CMECB induced Jurkat T and Wil-2 NS cell division cycle arrest at the G2/M phase as determined by flow cytometric analysis. Western blot analyses of G2/M phase regulatory proteins demonstrated that the CMECB-induced cell division cycle arrest was associated with the downregulation of cyclin B1 and Cdc2 protein expression levels. Western blot analyses results further revealed that the arrest of Wil-2 NS cells at the G2/M phase was independent of p21 protein activity. However, Jurkat T cell division cycle arrest was found to be mediated, in part, by p21. Quantitative real-time PCR results did not show a clear trend in terms of the down- or up-regulatory effects of the extracts on the G2/M phase regulatory genes. The CMECBinduced apoptosis and G2/M arrest was found to occur in a p53-independent xxii manner due to the lack and down-regulation of p53 protein levels in both Jurkat T and Wil-2 NS cells, respectively. In conclusion, CMECB induces its anticancer activity by inducing G2/M phase arrest and mitochondrial-mediated apoptosis independent of p53 protein activity. Although the study did not perform in vivo experiments to ascertain the efficacy of extracts of CMECB against specific tumour types in animal models, the present findings somehow validate the traditional use of C. benghalensis L as an anticancer agent. A more definitive study needs to be done to ascertain this assertion. / National Research Foundation and the University of Limpopo research office

MSc (Botany) / Department of Botany / See the attached abstract below

MSc (Botany)

MSc (Microbiology) / Department of Microbiology / See the attached abstract below.

MA (Tshivenda) / Senthara ya M. E.R. Mathivha ya Nyambo dza Afrika, Vhutsila na Mvelele / See the attached abstract below

MSc (Botany) / Department of Botany / Diabetes and arthritis are the most common chronic diseases. Arthritis is the leading cause of global disability and diabetes has become a major health problem which is increasing rapidly. The purpose of the study was to document medicinal plants that are used to treat and manage diabetes and arthritis by traditional medicinal practitioners around the Vhembe District Municipality as well as to evaluate their in vitro efficacy. Traditional practitioners were interviewed using semi-structured questionnaires. Seventeen plant species belonging to fourteen different families were found to be used in the treatment of diabetes as well as arthritis. Fabaceae family was dominating. Antioxidant, anti-inflammatory, cytotoxicity, alphaamylase and alpha-glucosidase) of five plant species, (Bridellia mollis, Elephantorrihiza burkei, Elaeodendron transvaalense, Senna petersiana and Searsia lancea) used traditionally to manage diabetes were investigated using the standard in vitro procedures. All extracts showed a good nitric oxide inhibition, with highest percentage inhibition found in the highest concentration of 100 μg/ml. They all had good percentage cell viability at lowest concentration which was comparable to quercetin. Only two plant extracts B. mollis (T2) and E. transvaalense (T3) had lower than inhibition of quercetin at 25 μg/ml than at 12.5 μg/ml. In vero cells low toxicity effect was observed at lowest concentration tested, and toxicity increased with the increase in concentration. In bovine dermis cell line all plant extracts had more cell viability at lower concentration than doxorubicin. Ethanol extracts of B. mollis and S. petersiana, and ethyl extract of E. transvaalense had a good alpha-amylase inhibitory activity with IC50 values 58.6, 81.9 and 131.5 mg/ml respectively. Hydro-ethanol, ethyl acetate and ethanol extracts of E. burkei exhibited a significant alpha-glucosidase inhibitory activity with IC50 values 56.9, 52.2 and 129.7 mg/ml respectively. Kinetic analysis revealed non-competitive and un-competitive inhibitions of the plant extracts on alpha-amylase and alpha-glucosidase enzymes respectively. The information obtained showed that people in Vhembe District Municipality still rely on medicinal plants to treat and manage diabetes and arthritis. All plant extracts were toxic to both bovine dermis and vero cell lines. S. lancea (T5) was found to be the most toxic plant extract. The observed good inhibitions of both alpha-amylase and alpha-glucosidase enzymes by plant extracts of B. mollis, S. petersiana, E. transvaalense and E. burkei validate their use in the traditional treatment of diabetes in the region to some extent. Aqueous extracts of these medicinal plants should also be investigated because water is the main solvent which is used by traditional practitioners in the preparation of their herbal medicines.

MSc (Botany) / Department of Botany / Understanding the uses of indigenous plants that are of economic importance to local communities is very much important in rural development strategies. The Marula (Sclerocarya birrea) Anacardiaceae family is widely used. More information on this tree species would enhance its value in agricultural landscapes, by helping farmers improve their livelihoods and ensuring environmental sustainability. Understanding how a community uses a resource and what influences the level of its use is crucial for developing a framework for its sustainable use based on local demands. Sclerocarya birrea is a species with multiple uses, which is recognized as commercially, medicinally and culturally important in Africa. Almost all parts of this species are useful. The study presented the findings of a survey of the indigenous knowledge, uses and management of S. birrea in Matshena village, Limpopo Province, South Africa. Different people of various ages were interviewed using a semi-structured questionnaire. Thirty percent of respondents indicated that they utilize the marula for beer and juice-making, the highest use category. In the sampled area the population of S. birrea is dominated by larger trees with no seedlings and juveniles. This is a sign of a population that will not be viable, since there are no younger individuals to replace the older trees when they die.

PhD (Microbiology) / Department of Microbiology / See the attached abstract below