A Simplified Liquid Chromatography-Mass Spectrometry Assay for Artesunate and Dihydroartemisinin, Its Metabolite, in Human Plasma
AbstractArtesunate (AS) is a potent antimalarial that is used worldwide for the treatment of malaria. A simple method with a total run time of 12 min was developed and validated for the quantification of AS and dihydroartemisinin (DHA), its active metabolite, in human (heparinized) plasma based on one-step protein precipitation in acetonitrile using artemisinin (ARN) as an internal standard, followed by liquid chromatography with a single quadrupole mass spectrometry system connected to a C18 column. Peak area ratio responses were fitted to the 2nd-order curve type, polynomial equation with weighting (1/concentration) over a quantification range between 3.20/5.33–3,000/5,000 nM (1.23/1.52–1153/1422 ng/mL) of AS/DHA showing linearity with very good correlation (r2 > 0.999). Single ion recordings of 5 µL injections of plasma extracts allowed for limits of detection of 1.02 nM (0.39 ng/mL) for AS and 0.44 nM (0.13 ng/mL) for DHA. The inter-assay and intra-assay accuracy and precision of the method was very good with an inaccuracy of ±12.4% and coefficients of variation of ≤10.7% at all tested concentrations. The recovery of the analytes from plasma was ≥95%. Other commonly used antimalarials including mefloquine, quinine, and chloroquine, did not interfere with the analysis. Post-preparative tests over 24 h in an autosampler (10 °C) showed that the DHA response was only 2.1% of AS from auto-hydrolysis, and β-DHA was the major, stable epimer that was used for quantification of DHA. In contrast, α-DHA increased steadily up to 600%. Artesunate and DHA in plasma were stable through three freeze/thaw cycles for up to 6 h at room temperature and up to one year at -80 °C.
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Teja-Isavadharm, P.; Siriyanonda, D.; Siripokasupkul, R.; Apinan, R.; Chanarat, N.; Lim, A.; Wannaying, S.; Saunders, D.; Fukuda, M.M.; Miller, R.S.; Weina, P.J.; Meléndez, V. A Simplified Liquid Chromatography-Mass Spectrometry Assay for Artesunate and Dihydroartemisinin, Its Metabolite, in Human Plasma. Molecules 2010, 15, 8747-8768.
Teja-Isavadharm P, Siriyanonda D, Siripokasupkul R, Apinan R, Chanarat N, Lim A, Wannaying S, Saunders D, Fukuda MM, Miller RS, Weina PJ, Meléndez V. A Simplified Liquid Chromatography-Mass Spectrometry Assay for Artesunate and Dihydroartemisinin, Its Metabolite, in Human Plasma. Molecules. 2010; 15(12):8747-8768.Chicago/Turabian Style
Teja-Isavadharm, Paktiya; Siriyanonda, Duangsuda; Siripokasupkul, Raveewan; Apinan, Roongnapa; Chanarat, Nitima; Lim, Apassorn; Wannaying, Srisombat; Saunders, David; Fukuda, Mark M.; Miller, Robert S.; Weina, Peter J.; Meléndez, Victor. 2010. "A Simplified Liquid Chromatography-Mass Spectrometry Assay for Artesunate and Dihydroartemisinin, Its Metabolite, in Human Plasma." Molecules 15, no. 12: 8747-8768.