Publication: High-performance liquid chromatographic assay for thymidylate synthase from the human malaria parasite, plasmodium falciparum
Issued Date
1989-01-01
Resource Type
ISSN
03784347
Other identifier(s)
2-s2.0-0024588482
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Mahidol University
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SCOPUS
Bibliographic Citation
Journal of Chromatography B: Biomedical Sciences and Applications. Vol.487, No.C (1989), 51-59
Suggested Citation
Jerapan Krungkrai, Yongyuth Yuthavong, H. Kyle Webster High-performance liquid chromatographic assay for thymidylate synthase from the human malaria parasite, plasmodium falciparum. Journal of Chromatography B: Biomedical Sciences and Applications. Vol.487, No.C (1989), 51-59. doi:10.1016/S0378-4347(00)83006-6 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/15730
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Title
High-performance liquid chromatographic assay for thymidylate synthase from the human malaria parasite, plasmodium falciparum
Abstract
A rapid and highly sensitive high-performance liquid chromatographic assay for thymidylate synthase activity is described. The assay is based on the separation of the substrate, deoxyuridylate (dUMP), and its product, deoxythymidylate (dTMP), on a LiChrosorb RP-8 reversed-phase column with 44 mM triethylammonium phosphate (pH 7.0) as mobile phase and a flow-rate of 1.0 ml/min. In addition, using a μBondapak C 18 reversed-phase column with 10 mM potassium phosphate (pH 4.0) and a gradient of 0-28% methanol, dUMP, dTMP and deoxythymidine (dTdR) are well separated within 30 min. The latter system is also applied to assay thymidine kinase activity with dTdR and dTMP as substrate and product, respectively. This method is sensitive enough to measure dTMP at concentrations as low as 25 pmol, and it was used to show that crude extracts of the human malaria parasite Plasmodium falciparum contain thymidylate synthase but not thymidine kinase activity. © 1989.