title: A colorimetric high-throughput beta-hematin inhibition screening assay for use in the search for antimalarial compounds.
abstract: Antimalarial drugs such as chloroquine are believed to act by inhibiting hemozoin formation in the food vacuole of the malaria parasite. We have developed a new assay for measuring and detecting inhibition of synthetic hemozoin (beta-hematin) formation. Aqueous pyridine (5% v/v, pH 7.5) forms a low-spin complex with hematin but not with beta-hematin. Its absorbance obeys Beer's law, making it useful for quantitating hematin concentration in hematin/beta-hematin mixtures, allowing compounds to be investigated for inhibition of beta-hematin formation. The assay is rapid (60 min incubation) and requires no centrifugation. The beta-hematin inhibition data show good agreement with alternative assay methods reported by four laboratories. The assay was adapted for high-throughput colorimetric screening, allowing visual identification of beta-hematin inhibitors. In this mode, the assay successfully detected all 18 beta-hematin inhibitors in a set of 47 compounds tested, with no false positive results. The quantitative in vitro antimalarial activities of a set of 13 aminoquinolines and quinoline methanols were found to correlate significantly with beta-hematin inhibition values determined using the assay.
authors.affiliation: Department of Chemistry, University of Cape Town, Private Bag, Rondebosch 7701, South Africa.
authors.forename: Kanyile K
authors.initials: KK
authors.lastname: Ncokazi
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Egan
mesh_terms: D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D002738:Chloroquine / Q000494:pharmacology; D003124:Colorimetry / Q000379:methods*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006420:Hemeproteins / Q000037:antagonists & inhibitors* / Q000737:chemistry; D020128:Inhibitory Concentration 50; D011725:Pyridines / Q000737:chemistry
chemical_list: D000962:Antimalarials; D006420:Hemeproteins; D011725:Pyridines; C020753:hemozoin; D002738:Chloroquine; C023666:pyridine