A colorimetric high-throughput β-hematin inhibition screening assay for use in the search for antimalarial compounds

被引:172
作者
Ncokazi, KK [1 ]
Egan, TJ [1 ]
机构
[1] Univ Cape Town, Dept Chem, ZA-7701 Rondebosch, South Africa
基金
新加坡国家研究基金会; 英国医学研究理事会;
关键词
D O I
10.1016/j.ab.2004.11.022
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
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% nu/nu, 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 P-hematin formation. The assay is rapid (60 min incubation) and requires no centrifugation. The P-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. (c) 2004 Elsevier Inc. All rights reserved.
引用
收藏
页码:306 / 319
页数:14
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