In Vitro Adventitious Regeneration of Artemisia annua L. Influencing Artemisinin Metabolism

Artemisia annua L. is a herbaceous plant belonging to the Asteraceae family, known for producing, although at low levels, the sesquiterpene lactone artemisinin (AN), which is highly effective against malaria. In this study, an in vitro regeneration process of A. annua L. using 'Artemis' progeny was established and the potential of tissue culture for inducing new variability in terms of AN metabolism of in vitro regenerated plants was investigated. Among the plant growth regulators tested, the cytokinin 6-benzyladenine (BA) at 4.4 mu M in combination with the auxin indole-butyric acid (IBA) at 0.35 mu M yielded the greatest frequency of shoot induction. The optimal multiplication medium contained BA at 0.9 mu M and naphthaleneacetic acid (NAA) at 0.05 mu M. Regenerated plants (RPs), after transferring to the greenhouse and subsequently to the field, were analyzed during the growth cycle at different sampling times, showing a peak of AN content 20 days before blossom. Variability among different RPs and sampling times, in terms of AN and its precursors dihydroartemisinic acid (DHAA) and artemisinic acid (AA) was observed. This suggests that adventitious shoot induction could provide a useful strategy to induce variability influencing artemisinin metabolism as a consequence of in vitro manipulation.