Enzymatic Synthesis of Artemisinin from Natural and Synthetic Precursors

Abstract

To investigate the biosynthetic pathway of artemisinin, an assay system for the determination of activity of the enzymes involved in its synthesis has been developed. Results from these experiments have shown that HEPES provides a better buffer system than Tris-HCl. The enzyme(s) requires Mg2+ and/or Mn2+, and the addition of ATP and NADPH+H+ significantly enhances the enzyme activity. A new substrate, dihydroarteannuin B, has been synthesized that can easily be radiolabeled with high specific activity. It is utilized by the enzyme system and is converted to artemisinin with the same efficiency as the natural substrates. This can be conveniently used as a precursor for elucidation of the pathway for artemisinin biosynthesis.

Quantification of Three Key Enzymes Involved in Artemisinin Biogenesis in Artemisia annua by Polyclonal Antisera-Based ELISA

“…Subsequent experiments by others using various precursors and intermediates agreed with this conclusion (Sangwan et al 1993;Nair and Basile 1993;Bharel et al 1998;Dhingra et al 2000;Dhingra and Narusu 2001). More recently, however, the work reported by Brown and Sy (2007) strongly supports an alternative hypothesis that proposes dihydroartemisinic acid rather than artemisinic acid as the immediate precursor of artemisinin ).…”

In vivo transformations of dihydroartemisinic acid in Artemisia annua plants

“…(It should be noted that compound 3 is also hydroxylated and derivatized as glycosides by plant tissue cultures of A. annua, in what appears to be a degradative pathway.) 16 For example, while both Nair and Basile 23,24 and Jain et al 25 have found experimental evidence that arteannuin B (4), or its 11,13-dihydro analogue, dihydroarteannuin B (5), can be converted into artemisinin (1) by cell-free extracts, Wang et al have noted that compound 4 was not a precursor of 1, 15 but suggested rather that epi-deoxyarteannuin B (6) and dihydro-epi-deoxyarteannuin B (7) are the true intermediates in the conversion of 2/3 to 1. 26 They also rejected the 11,13-dehydro natural product, a-epoxy-artemisinic acid (8), 27 as a biogenetic precursor of 1, 15 and it has been proposed by these (and other) authors that the 11,13-dehydro-amorphane artemistene (9) may, in fact, be the immediate precursor of 1.…”

A golden phoenix arising from the herbal nest — A review and reflection on the study of antimalarial drug Qinghaosu

“…Furthermore, on the incubation with the cell free extracts of Qinghao leaves, artemisinic acid (11) could also be transferred into 3, and in turn, 3 as well as dihydroarteannuin B (23) could be transferred to QHS. Therefore, they concluded that 3 was the biogenesis precursor of QHS, but it has not been mentioned whether 3 is the necessary intermediate for the biosynthesis of QHS [154][155][156]. It is unclear whether the different conclusions about the role of arteannuin B (3) came from the different experimental conditions, isotopically labeled precursor in the homogenate [152] versus unlabeled precursor in a cell free system [155].…”