Exploring the peroxidase activity of ferriprotoporphyrin IX: towards understanding its cytotoxicity in the malaria parasite
| dc.contributor.advisor | De Villiers, Katherine A. | en_ZA |
| dc.contributor.advisor | Gerber, Wilhelmus J. | en_ZA |
| dc.contributor.author | Bergh, Wikus | en_ZA |
| dc.contributor.other | Stellenbosch University. Faculty of Science. Dept. of Chemistry and Polymer Science. | en_ZA |
| dc.date.accessioned | 2019-02-26T10:18:29Z | |
| dc.date.accessioned | 2019-04-17T08:06:16Z | |
| dc.date.available | 2021-02-28T03:00:09Z | |
| dc.date.issued | 2019-03 | |
| dc.description | Thesis (MSc)--Stellenbosch University, 2019. | en_ZA |
| dc.description.abstract | ENGLISH ABSTRACT: The peroxidase reaction, catalysed by ferriprotoporphyrin IX (Fe(III)PPIX), was selected as a model reaction by which to explore Fe(III)PPIX toxicity via the formation of reactive oxygen species. This has direct relevant in, for example, antimalarial chemotherapy. To analyse potential reaction steps, model-based analysis is used. A program was coded incorporating a simplex algorithm for the multivariate global analysis of experimental data using non-linear least squares (NLLS) regression analysis. The reaction between Fe(III)PPIX and hydrogen peroxide was studied, with tests conducted regarding reversibility of proposed reaction steps, as well as the addition of potential side reactions. NLLS regression analysis of proposed models yielded fitted data that could be used to determine the goodness-of-fit of the model, as well as compare fits of models as a function of their complexity. Model comparison analysis showed that reversibility of reactions were irrelevant, and that the best depiction of the reaction proceeds via three steps. The first is complexation of Fe(III)PPIX and H2O2. Secondly, the complex forms an active intermediate 𝐼. The last step is denoted by the conversion of the intermediate to Fe•+(IV)PPIX=O. Degradation of Fe•+(IV)PPIX=O was incorporated in model extensions as side reaction but did not form part of the model determined to be best. ABTS is used as chromogen to follow the peroxidase-like reaction of Fe(III)PPIX spectrophotometrically. Studies conducted on plausible side reactions of the chromogen ABTS showed that the compound does not react with hydrogen peroxide at pH levels of 4.8 and 7.5. The disproportionation reaction is shown to occur under highly acidic conditions but is concluded not to transpire at pH levels approaching physiologically relevant conditions. Degradation of ABTS is not observed for physiologically relevant conditions. ABTS•+ degradation is considered a viable side reaction under the premise that it can be reduced to a null parameter following non-linear least squares regression in the case that it does not prove relevant. Results obtained for the reaction between Fe(III)PPIX and H2O2 and ABTS studies are incorporated in the proposition of plausible reaction models for the peroxidase-like reaction of Fe(III)PPIX. NLLS analysis of models presented based on literary review indicate that the peroxidase-like activity of Fe(III)PPIX involves two parallel catalytic cycles. One involves the reaction steps presented for the reaction of Fe(III)PPIX with H2O2. From the formation of Fe•+(IV)PPIX=O, the radical porphyrin compound can react with ABTS to form Fe(IV)PPIX=O and ABTS•+. Fe(IV)PPIX=O can react with another molecule of ABTS to form ABTS•+ and regenerate Fe(III)PPIX, thus concluding the cycle. The second catalytic cycle is initiated by the complexation of Fe(III)PPIX with ABTS. The complex formed can react with H2O2 to produce a Fe•+(IV)PPIX=O.ABTS complex. The porphyrin radical compound can react with the ABTS molecule associated to it to produce ABTS•+ and Fe(IV)PPIX=O, thus joining into the first cycle presented. The model proposed can be used to compare the kinetics of this reaction in the presence of antimalarial compounds to elicit the effect these compounds have on the toxicity of Fe(III)PPIX; such insights may advance rational design efforts towards new antimalarial chemotherapeutics. | en_ZA |
| dc.description.abstract | AFRIKAANSE OPSOMMING: Die peroksidase reaksie, gekataliseer deur ferriprotoporfirien IX (Fe(III)PPIX), was gekies as ‘n model reaksie om die toksisiteit van Fe(III)PPIX wat reaktiewe suurstof spesies form te bestudeer. Hierdie is direk relevant vir byvoorbeeld antimalariese chemoterapie. Om potensiele reaksie stappe te analiseer is model-gebaseerde analise gebruik. ‘n Program was gekodeer wat ‘n simpleks algoritme inkorporeer vir die multivariate globale analise van experimentele data deur gebruik te maak van nie-lineêre kleinste vierkant regressie analise (NLKV). Die reaksie tussen Fe(III)PPIX en waterstof peroksied was bestudeer, met toetse wat die omkeerbaarheid van die voorgestelde reaksie stappe sowel as potensiele sy-reaksies in ag te neem. NLKV regressie analise van die voorgestelde modelle was gebruik om te bepaal hoe goed die model pas, asook om die passing van die model tot sy kompleksiteit te vergelyk. Model vergelykings analise het die omkeerbaarheid van sekere reaksies as irrelevant aangetoon, en verder gewys dat die beste beskrywing van die kinetiese reaksies volgens drie stappe plaasvind. Die eerste reaksie betrek die kompleksasie van Fe(III)PPIX en H2O2. Tweedens form die kompleks ‘n aktiewe intermediêre spesies, 𝐼. Die laaste stap word aangetoon deur die omskakeling van die intermediêre spesies na Fe•+(IV)PPIX=O. Degredasie van Fe•+(IV)PPIX=O was in model verlengings geinkorporeer as sy-reaksies, maar het nie deel gevorm van die model meganisme wat die beste passings toon nie. ABTS word as chromogeen gebruik om die peroksidase-tipe reaksie van Fe(III)PPIX spektrofotometries te bestudeer. Studies gedoen om die moontlike reaksies van die chromogeen ABTS te bepaal wys dat die verbinding nie met H2O2 reageer by pH vlakke van 4.8 en 7.5 nie. Die disproporsionasie reaksie vind net plaas onder hoogs suur kondisies en dus nie by pH vlakke naby aan fisiologiese relevante kondisies nie. Degredasie van ABTS is nie waargeneem by fisiologiese relevante kondisies nie. ABTS•+ degredasie is oorweeg as ‘n potensiêle reaksie onder die premie dat dit gereduseer kan word tot ‘n nul parameter tydens die NLKV regressie analise van voorgestelde modelle in die geval wat die reaksie nie relevant is nie. Resultate verkryg vir die reaksies tussen Fe(III)PPIX en H2O2 asook ABTS studies is geinkorporeer in die voorstelling van moontlike reaksie modelle vir die peroksidase-tipe aktiwiteit van Fe(III)PPIX. NLKV regressie analise van modelle gebaseer op literatuur wys dat die peroksidasie-tipe aktiwiteit van Fe(III)PPIX twee paralelle katalitiese siklusse bevat. Die hoof siklus bevat die reaksie stappe soos bepaal vir die reaksie van Fe(III)PPIX met H2O2. Die gevormde van Fe•+(IV)PPIX=O radikale porfirien verbinding reageer dan met ABTS om Fe(IV)PPIX=O en ABTS•+ te vorm. Fe(IV)PPIX=O kan ook reageer met nog ‘n molekule van ABTS om ABTS•+ te vorm en Fe(III)PPIX te hergenereer, en so die eerste siklus te voltooi. Die tweede katalitiese siklus word geinisieer deur die kompleksasie van Fe(III)PPIX met ABTS. Die gevormde kompleks kan met waterstofperoksied reageer om ‘n Fe•+(IV)PPIX=O.ABTS kompleks te produseer. Die porfirien radikaal verbinding kan met die ABTS molekule aan hom verbind reageer om ABTS•+ en Fe(IV)PPIX=O te produseer, en dus by die hoof siklus aan te sluit. Die voorgestelde model kan gebruik word om die kinetika van die reaksies in die teenwoordigheid van antimalariese dwelms te vergelyk, sodat die effek van hierdie dwelms op die toksisiteit van Fe(III)PPIX ontleed kan word. Sulke insigte mag dalk help in die rasionele ontwikkeling van nuwe antimalariese middels. | af_ZA |
| dc.description.version | Masters | en_ZA |
| dc.embargo.terms | 2021-02-28 | |
| dc.identifier.uri | http://hdl.handle.net/10019.1/105642 | |
| dc.language.iso | en_ZA | en_ZA |
| dc.publisher | Stellenbosch : Stellenbosch University | en_ZA |
| dc.rights.holder | Stellenbosch University | en_ZA |
| dc.subject | Heme oxygenase | en_ZA |
| dc.subject | Haem oxygenase | en_ZA |
| dc.subject | Hydrogen Peroxide | en_ZA |
| dc.subject | Peroxidase | en_ZA |
| dc.subject | ABTS | en_ZA |
| dc.subject | Model-based analysis | en_ZA |
| dc.title | Exploring the peroxidase activity of ferriprotoporphyrin IX: towards understanding its cytotoxicity in the malaria parasite | en_ZA |
| dc.type | Thesis | en_ZA |