| Science Letters |
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| Affinity ultrafiltration of DNA topoisomerases-targeted compounds determined with HPLC/ESI-MS for drug candidate screening |
| ZHANG Hong, PAN Yuan-jiang |
| Department of Chemistry, Zhejiang University, Hangzhou 310027, China; Department of Food Science, Hangzhou University of Commerce, Hangzhou 310035, China |
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Abstract A method of screening assay is demonstrated. The approach is based on the affinity of antitumor candidates for topoi-somerases. In this method, antitumor candidates are fished out using topoisomerases as targets. Traditional analysis of complex compounds typically encounters signal suppression due to the relatively low concentrations, but enzyme-affinity screening for the active compounds can effectively concentrate the desired analysts into a small volume of high concentration. Active compounds are separated from non-affinity compounds by ultrafiltration. The molecules-enzymes complexes that are retained on the filter are subsequently separated by acidification to obtain the topoisomerases-affinity compounds for analysis on High Performance Liquid Chromatography coupled with electrospray ionization mass spectrometric detection (ESI-MS). This enzyme-affinity based screening assay provides a highly specific and efficient method that can directly screen, identify, and acquire drug candidates thus improving the accuracy and speed of high-throughput screening activities.
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Received: 02 March 2004
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