Interactions of ferriprotoporphyrin IX with neutral lipids and detergents: insights into their role in β-haematin formation

Doctoral Thesis


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University of Cape Town

The malaria parasite ingests between 80-100% of the host red blood cell's iron in the form of haemoglobin, which is catabolised to amino acids and ferriprotoporphyrin (Fe(III)PPIX), that it subsequently detoxifies to form the bio crystal, haemozoin. Neutral lipids have been implicated as the biological molecules responsible for the nucleation of haemozoin in vivo, though their exact role is unclear. This thesis has investigated the interaction between these lipids and Fe(III)PPIX. By exploiting the fluorescence quenching ability of Fe(III)PPIX, Stern-Volmer plots were generated to estimate the concentration of Fe(III)PPIX partitioned into Nile red stained neutral lipid droplets (NLBDs) over a pH range. The pH dependence was found to correlate with the charge of the Fe(III)PPIX molecule and the largest amount of Fe(III)PPIX partitioning was observed under the acidic pH conditions of the parasite digestive vacuole (pH 4.80), where haemozoin is formed. From the fluorescence maximum of Nile red, the relative polarity inside the lipid droplets was shown to lie between that of acetone and octanol with a Dimroth-Reichart ET(30) parameter of 45 kcal/mol. The lipophilicity of Fe(III)PPIX was validated by measuring the octanol-water partition coefficient, log DOW, as1.8 for the ionised form at pH 7.5 and a log P of 2.8 for the neutral form. 4-aminoquinoline drugs, chloroquine and amodiaquine, decreased the partitioning of Fe(III)PPIX into NLBDs at low pH owing to the consistently charged nature of the drug-Fe(III)PPIX complex. In contrast, the quinoline methanols, quinidine and quinine, strongly increased the observed lipophilicityof Fe(III)PPIX in a pH dependent manner. Quinidine was found to effect the largest increase eon Fe(III)PPIX partitioning into NLBDs which was proposed to be due to a previously established inherent increased lipophilicity of the quinidine-Fe(III)PPIX complex.

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