Effect of dimethyl-beta-ciclodextrins on artemisinin production in Artemisia annua suspension cell culture.

Artemisinin is an effective antimalarial compound obtained by thè aerial parts of thè herbal plant Artemisia annua L. Unfortunately, thè production of artemisinin in plants is very low (0.1 -1% on a dry weight basis) and its chemical synthesis is very difficult. In recent years biotechnological attempts to improve artemisinin production either in plant or genetically engineered microrganisms were made. A. annua celi and tissue cultures were also explored for thè production of artemisinin, although thè yieids obtained have been so far quite low. We have recently established A. annua celi cultures able to biosynthesize artemisinin and to respond to thè elicitation by methyl jasmonate (MeJA). Interestingly, amounts of artemisinin produced by these cultures were also found in thè spent culture medium. Cyclodextrins (CDs) are known to increase thè water solubility of various compounds, by forming host-guest inclusion complexes. Chemically modified CDs, such as 2,6 dimethyl-p-cyclodextrins (DIMEB) have been successfully used to improve thè production of health-promoting plant metabolites such as resveratrol in grape celi cultures. The aim of this work was to evaluate thè ability of DIMEB to enhance artemisinin production in A. annua suspension celi cultures. HPLC analysis of artemisinin and Real Time PCR gene expression analysis were carried out in DIMEB treated and untreated cultures. DIMEB induced a 300-fold increase of artemisinin levels in thè spent culture medium after a three-day-treatment. The addition of MeJA induced a further increase of thè artemisinin amounts. The expression levels of artemisinin biosynthetic genes (cytochrome P450 monoxygenase, cytochrome P450 reductase and artemisinic aldehyde A11 reductase) were not altered by thè DIMEB treatment. These results suggest that DIMEB improved artemisinin yieids of A. annua celi cultures, probably not affecting thè biosynthetic pathway but enhancing thè solubility and stability of artemisinin molecules.