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Analysis of underivatized artemisinin and related sesquiterpene lactones by high-performance liquid chromatography
Jorge F S Ferreira1, Javier M Gonzalez
1Appalachian Farming Systems Research Center, U.S. Department of Agriculture Agricultural Research Service, 1224 Airport Road, Beaver, WV 25813, USA. jorge.ferreira@ars.usda.gov
Introduction:
Although high-performance liquid chromatography with ultraviolet detection (HPLC-PAD) is widely available, it has not been used for artemisinin (1) analysis because of the lack of UV absorption reported for this lactone. Increased Artemisia annua cultivation for production of 1 requires an affordable and reliable method to analyse 1 and its precursors dihydroartemisinic acid (2) and artemisinic acid (3) simultaneously from underivatized plant extracts.
Objective:
To validate HPLC-PAD for the quantification of underivatized artemisinin from A. annua and artemisinin-based drugs.
Methodology:
Dried A. annua leaves were extracted with petroleum ether, dried, reconstituted in acetonitrile, and analysed by HPLC-PAD at 192 nm using an isocratic mobile phase (60:40, acetonitrile:0.1% acetic acid). HPLC-PAD was evaluated through accuracy, precision, recovery and comparison with HPLC with evaporative light scattering detection (HPLC-ELSD).
Results:
HPLC-PAD proved accurate, precise and reproducible for the direct quantification of 1 and related compounds, and was more sensitive than ELSD for most of the compounds tested. The limit of quantification of 1-3 from plants was 0.048, 0.024 and 0.008 g/100 g dry weight, respectively. Recoveries were over 98%, with good intra- and inter-day repeatability. HPLC-PAD correlated significantly (r(2 )= 0.99, p < 0.001) with HPLC-ELSD for artemisinin analysis. HPLC-PAD was also reliable for the analysis of dihydroartemisinin, artesunate and artelinic acid.
Conclusion:
HPLC with ultraviolet detection was validated for the quantification of underivatized 1, 2, and 3 from crude plant samples, and is readily applicable for the quality control of herbals and artemisinin-related pharmaceutical compounds.
