Morphological Study Of In Vitro Culture Of Artemisia Annua L. And Enhancement Of Artemisinin Production Via Chemical Mutagenesis
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Date
2014-07
Authors
Leow, E Shuen
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Publisher
Universiti Sains Malaysia
Abstract
Artemisia annua L. plant is the source of production of artemisinin. Artemisinin was proven to be the most effective anti-malarial drug against multi-drug resistant Plasmodium spp. However, artemisinin content is relatively low in A. annua L. plant. Therefore, chemical mutagenesis using sodium azide (NaN3) and ethyl methanesulfonate (EMS) were incorporated in order to create variability with the objective of increasing the artemisinin content. The potential mutagenic effect of NaN3 and EMS was studied on seeds, explants from shoot apical meristem, nodal segments and callus of A. annua L. clones of Vietnam origin (TC1, TC2 and Highland) for the morphological changes and enhancement of artemisinin content. Results showed that a small percentage of putative mutant plant from NaN3 and EMS treated seeds, shoot tips and nodal segments had shown morphological abnormalities during the first two subculture cycles. However, these abnormalities subsided through each subsequent subculture cycle. The height, the leaf size and the internode length of putative mutant plants were recorded and no significant difference were observed after four subculture cycles. Nonetheless, the glandular trichomes on both sides of the A. annua L. leaves of the three tested clones had shown increased in density and/or size of glandular trichome after treatment with different concentration of NaN3 and EMS. The glandular trichomes are believed to be the major and possibly the solitary site for biosynthesis and accumulation of artemisinin. The increased in number of glandular trichomes was found to increase the artemisinin content in putative mutant plantlets. The artemisinin content in callus treated with NaN3 and EMS was not significantly increased. Eight new A. annua L. varieties were selected based on the size and density of glandular trichome and artemisinin content quantified using high-performance liquid chromatography (HPLC). The mutant plants with dark green leaves and deeply dissected lobes were found to have high trichome density and this indirectly is an indication of high artemisinin content. In conclusion, both NaN3 and EMS mutagenesis on seeds, shoot tips and nodal segments were suitable alternatives for creating variability for producing new varieties in A. annua L. with enhanced artemisinin. However, results shown in EMS mutagenesis were more consistent in general as compared to NaN3 mutagen. For effective and successful mutagenesis of A. annua L, the optimum NaN3 concentration was set at 0.06 - 0.1mM whilst the optimum concentration for EMS was 1.00% (v/v).
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Keywords
Artemisia annua L. plant , source of production of artemisinin