Artemisinin (Qinghaosu) is a potent antimalarial produced by the Chinese herb, Artemisia annua L. and is particularly effective in the treatment of the parasitic infections caused by chloroquine-resistant Plasmodium species. In clinical studies conducted in the Peoples Republic of China, spanning over a decade, involving thousands of human patients, artemisinin was found to be relatively safe and free of adverse side effects. However, at present, it is available for treatment, only in the Peoples Republic of China where the independent clinical evaluation as a prophylactic, and for mass treatment in other areas where malaria is endemic, is particularly urgent now, because of the steady proliferation of chloroquine-resistant strains. The primary objective of this project is to employ, as complementary, or as an alternative, to whole plant extraction, the following highly effective biotechnological approaches, namely, cell suspension cultures and bioconversion using cell-free enzymes. Dr. Nair and co-workers recently demonstrated in preliminary experiments that cell suspension cultures of A. annua produces artemisinin. They now propose to obtain new cell lines which are better producers of the drug, by varying the growth hormones and other parameters governing callus formation. These cell lines will then be used in suspension cultures under varying conditions to optimize artemisinin production. Finally, pilot plant scale studies will be initiated to determine the economic viability of the process. Intermediates in the biosynthesis of a compound can be less complex and thus more easily accessible by synthesis, and hence, are attractive starting compounds for bioconversion studies. Terpenoids closely related to artemisinin, as well as those of cadinane and germacrane groups, have been postulated as precursors of it. However, the only reported biotransformation using cell-free extracts has been that of arteannuin B. Now, artemesic acid, another metabolite of A. annua, which is more abundant (> 10 times) than artemisinin in some strains, has also been converted to the latter. Parallel to these studies, they will use labeled, basic building blocks to define the biogenetic pathway of this terpenoid and to identify the intermediates. Partially purified and/or immobilized enzymes derived for leaves and cell cultures will be used for bioconversions. The proposed approaches will lead to additional routes for an adequate supply of artemisinin; they also can result in the isolation of more potent derivatives of the drug by its bioconversion by the cell cultures and cell-free extracts.
