Masters Degrees (Pharmacology)

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Browsing Masters Degrees (Pharmacology) by Author "Du Plessis, J. B."

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    The effect of Phela, a traditional medicine on the immune system of a rat model
    (University of the Free State, 2010-08) Lekhooa, 'Makhotso Rose; Walubo, A.; Du Plessis, J. B.; Matsabisa, M. G.
    English: Phela is a herbal traditional medicine product prepared using well defined parts of four African medicinal plants. The aim of the study was to determine the mechanism of action by which phela boosted the immune system of a rat model. Unfortunately, subsequent literature research revealed that very little is known about phela. As such chromatographic methods were developed by which to identify phela and for quality control purposes. Of note, the developed methods complemented each other; they were compiled into a comprehensive method for phela fingerprinting. It involved sample extraction of the phela by either acidic extraction or a simple “saltingout” method, followed by Thin Layer Chromatography (TLC), and/or preparative Column Chromatography (CC) that were supported by High Performance Liquid Chromatography with UV-detector (HPLC_UV), HPLC with fluorescence detector (HPLC_FL), HPLC with photo diode array detector (HPLC_PDA) and Gas Chromatography-Mass selective detector (GC_MSD) spectrometry. The method was successfully used to differentiate phela from another herbal product made from Hypericum perforatum (St John’s wort) illustrating its high potential for application to other herbal medicines in development. Thereafter a HPLC_UV method for monitoring phela in plasma was developed and validated. However, due to its pitfalls, a HPLC_FL method was developed as well. The HPLC_UV method had a linear regression equation of y = 0.02x – 0.59, correlation coefficient of r2 = 0.9983 and accuracy within 15 %. For HPLC_FL three marker peaks were selected and standardized by the retention time. The developed HPLC_UV and HPLC_FL methods were tested by analysing blood collected from rats treated with phela after 1, 2, 4, 6 and 8 hours respectively. Unfortunately, analysis by HPLC_UV method had no peaks whereas, during HPLC_FL method analysis, a new peak was observed at 9.2 minutes. The peak was depicted as a metabolite for phela and was then utilized as the plasma marker. Since the metabolite is unknown and therefore no standard exists by which to express its plasma concentration in conventional units, the changes in peak area per unit volume (L) of plasma (peak-area/L) were used to derive the appropriate pharmacokinetic parameters hence peak-kinetics. The predictive parameters show that concentration of the metabolite at steady state would be 48 peak-area/ml, hence no accumulation of the drug. The final part of the study was undertaken to understand the mechanism of action of phela using a rat model by observing for changes in the levels of TH1 and TH2 cytokines. Rats were divided into six groups. The first group was not treated with anything. The four groups were treated daily with either normal-saline, orally (control); cyclosporine in olive oil, subcutaneously (negative control); Phela, orally (test-group 1) and phela+cyclosporine (test-group 2). The last group was inoculated once with influenza vaccine. The treatment period was 14 days and in each group rats were sacrificed after 7 and 14 days. Haematology tests, biochemistry tests, and cyclosporine level analysis were done. Serum cytokine analysis for TH1 (IL-2, IFN-γ and TNF-α) and TH2 (IL-4 and IL-10) cytokines was measured by ELISA. On days 7 and 14, the concentrations of TH1 cytokines in the Phela-only treated group were similar to the control. However, the TH1 cytokines were higher in the Phela+cyclosporine-A treated group than in the cyclosporine-A group, and cyclosporine- A concentrations were similar in both groups. These results show that Phela did not affect TH1 cytokines of a normal immune system but stimulated them when the immune system was suppressed by cyclosporine-A. This implies that Phela is a cyclosporine-A antagonist that may stimulate a compromised immune system. In conclusion, Phela is an immune stimulant.