Analysis of Artemisinin and Related Sesquiterpenoids fromArtemisia annuaby Combined Gas Chromatography-Mass Spectrometry

Abstract

Arteinisia annua L. (Asteraceae) is a source of the novel antimalarial drug artemisinin (1), that is active against Plas,nodiui'n parasites, including chioroquine-resistant strains, and effective in the treatment of cerebral malaria (1). For a better understanding of the artemisinin biosynthesis, knowledge of precursor contents and distribution in plant material is important.We developed a GC method, generally applicable on plant material, for the analysis of I and its precursors arteannuic acid (2) and arteannuin B (3) in one run. The method was validated by GC-MS, applied on pure reference standards (as solid probes and in methanolic solution) and extracts of different plant parts. cH3 H3C H2 2 cH3 H30 3The leaves, buds, and small stems contained the highest levels of 1, 2, and 3. Maximal concentrations were found in the buds: 0.29% 1, 0.68% 2, and 0.26% 3, calculated on DW. The main stem only contained minor amounts. In the side roots, 2 and 3 were found in concentrations of 0.017% and 0.006% DW, respectively, while 1 was absent. The main root was devoid of the three sesquiterpenoids.The GC method for the simultaneous determination of I and its precursors will be useful for the selection of highproducing plants, next to the semi-quantitative TLC analysis for artemisiriin (2), and is valuable for plant biotechnological purposes, to monitor the formation of secondary metabulites in in vitro cultured cells or tissues (3).

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“…A rapid screening method based on tandem mass spectrometry (MS/MS) is described for artemisinin-related compounds present in a crude hexane extract of A. annua [64]. Also, a detection method based on GC combined with MS [10,91,92] and radioimmunoassay or RIA have been described [97]. In 1993, Jaziri et al [34] reported the establishment of an indirect enzyme-linked immunosorbent assay (ELISA) method based on the peroxide bridge for antibody specificity to detect artemisinin and closely related compounds in crude extracts of A. annua.…”

“…Little attention has been paid to the analysis of late precursors in artemisinin biosynthesis, namely the sesquiterpenoids arteannuic acid, arteannuin B and artemisitene. TLC has been used for the simultaneous detection of these compounds, but the compounds are not fully resolved [91]. However, a method for the simultaneous determination of artemisinin-related compounds may be valuable for plant biotechnological purposes.…”

“…The formation of secondary metabolites in the in vitro cultured cells or tissues may then be monitored more appropriately. A GC/MS method for the analysis of artemisinin and its precursors arteannuic acid, arteannuin B and artemisitene in one run was developed [91]. Because of their thermal instability, the endoperoxide containing sesquiterpene lactones artemisitene and artemisinin were measured as their pyrolysis products.…”

“…Some disagreement exists about when the highest artemisinin content occurs. Some researchers state that the artemisinin content is highest just before flowering [1,26,48,91,92]. Others find an artemisinin peak reached during full flowering [28,54,62,75].…”

Determination of artemisinin in Artemisia species by hollow fiber-based liquid-phase microextraction and electrospray ionization-ion mobility spectrometry

“…High performance liquid chromatography, with different detection modes, such as UV, 4,5 electrochemical, 6 evaporative light scattering, 7 chemiluminescence 8 and mass spectrometric detection 9,10 is the most common technique. Capillary electrophoresis, 11 and gas chromatography (GC) 12,13 have also been used.…”