Browsing by Author "Sammy, Chandre Jade"
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- ItemInvestigations into the Peroxidase Activity of Ferriprotoporphyrin IX and its Complexes with Clinically Relevant Antimalarial Drugs(Stellenbosch : Stellenbosch University, 2017-03) Sammy, Chandre Jade; De Villiers, Katherine A.; Kuter, David; Stellenbosch University. Faculty of Science. Dept. of Chemistry and Polymer Science.ENGLISH ABSTRACT: A major class of antimalarial drugs act by increasing levels of free haem, Fe(III)PPIX, within the malaria parasite. While Fe(III)PPIX is known to be toxic, the manner in which it brings about parasite death is not entirely clear. Furthermore, it is unknown as to whether the antimalarial drugs which interact with Fe(III)PPIX, modulate its toxicity. The work presented here aims to address these uncertainties. The first part of this work focusses on the interactions between clinically relevant antimalarial drugs and Fe(III)PPIX. Spectrophotometric titrations were used to investigate drug-Fe(III)PPIX complex formation under three biologically relevant environments, namely an aqueous environment (pH 7.4) representative of the malaria parasite cytosol; acetonitrile which mimics the non-aqueous interior of a lipid and the detergent SDS was used to model a lipid-water interface system, that is present at membrane surfaces. Three antimalarial drugs chloroquine (CQ), quinidine (QD) and artemisinin (Art), belonging to three distinct classes, were investigated to determine binding strength between the antimalarial drug and Fe(III)PPIX. Strong association constants for the interaction of Fe(III)PPIX with QD and CQ were determined in the aqueous (log K = 5.8 ± 0.1 and 6.5 ± 0.1), aqueous SDS system (log K = 6.2 ± 0.1 and 6.5 ± 0.1) and acetonitrile (log K = 5.92 ± 0.1 and 5.3 ± 0.2). An association constant for the interaction of Fe(III)PPIX with Art could, however, only be determined in acetonitrile (log K = 4.48 ± 0.04) owing to a weakened interaction in aqueous media. Furthermore, the first crystal structure of the complex formed between Fe(III)PPIX and the QD analogue, cinchonine, was obtained following slow evaporation from a solution of acetonitrile. To probe the possible mechanism of toxicity of Fe(III)PPIX, its peroxidase activity was investigated using the chromogenic substrate, 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). The oxidation of ABTS to ABTS˙+ by Fe(III)PPIX and hydrogen peroxide (H2O2) was investigated in an aqueous and aqueous SDS solvent system. Radical production was found to be more stable in the aqueous SDS system, and furthermore, the rate of radical production was also found to be lower in this system compared to aqueous solution. From the available experimental data, attempts were made to determine a preliminary kinetic model for the oxidation of ABTS by Fe(III)PPIX and H2O2 in aqueous solution. The modulatory effect of CQ, QD and Ar on the oxidation of ABTS by Fe(III)PPIX was monitored in both aqueous solution and aqueous SDS. Both CQ and QD were found to significantly inhibit the rate of radical production in aqueous solution by 30% and 80%, respectively. While the rate of ABTS oxidation is lower, CQ was found to extend the life of Fe(III)PPIX by reducing the rate of its degradation by H2O2 by 70%, while QD reduced the rate of degradative attack by 90%. Consequently, Fe(III)PPIX has reduced peroxidative activity but is present for a longer period. The same was true for QD in the aqueous SDS system, however, CQ was found to have minimal effect on the yield of radical production compared to Fe(III)PPIX in aqueous SDS, however, the rate was still reduced. Art was found to have the opposite effect of CQ and QD and enhanced radical production by three times that of Fe(III)PPIX. This is thought to be related to the presence of the endoperoxide bridge in the structure of Art and it is proposed that the induction of oxidative stress is one of its possible mechanisms of action. The results obtained in this study provide insight into the mechanism of drug-Fe(III)PPIX interaction. Furthermore, valuable information regarding the ABTS oxidation catalytic cycle was determined through careful consideration of the kinetic model and the assay could be used to provide insight into the mechanism of toxicity of CQ, QD and Ar.