A colorimetric high-throughput β-hematin inhibition screening assay for use in the search for antimalarial compounds
A colorimetric high-throughput β-hematin inhibition screening assay for use in the search for antimalarial compounds
复制标题
DOI：
10.1016/j.ab.2004.11.022
复制
发表时间：
2005-03-15
影响因子：
2.9
通讯作者：
Egan, TJ
中科院分区：
文献类型：
--
作者：
Ncokazi, KK;Egan, TJ
抗疟疾药物如氯喹被认为是通过抑制疟原虫食物泡中疟原虫色素的形成而起作用的。我们已经开发了一种新的测定方法，用于测量和检测合成的血红素（β-血红素）形成的抑制。吡啶水溶液（5%nu/nu，pH 7.5）与血红素形成低自旋复合物，但不与β-血红素形成复合物。其吸光度符合比尔定律，可用于定量血红素/β-血红素混合物中的血红素浓度，从而研究化合物对β-血红素形成的抑制作用。该测定快速（孵育60分钟）且无需离心。P-血红素抑制数据显示与四个实验室报告的替代测定方法具有良好的一致性。该试验适用于高通量比色筛选，允许目视鉴定β-血红素抑制剂。在这种模式下，该检测方法成功检测了一组47种化合物中的所有18种β-血红素抑制剂，没有假阳性结果。一组13个氨基喹啉和喹啉甲醇的体外抗疟活性的定量被发现与使用该测定确定的β-血红素抑制值显着相关。(c)2004年爱思唯尔公司All rights reserved.
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.