Seasonal Variation of Artemisinin and its Biosynthetic Precursors in Plants of Artemisia annua of Different Geographical Origin: Proof for the Existence of Chemotypes

Abstract

The time course of the levels of artemisinin, its biosynthetic precursors and the biosynthetically related sesquiterpenes was monitored during a vegetation period of Artemisia annua plants of different geographical origin. Considerable differences in contents of artemisinin and its direct precursors artemisinic acid and dihydroartemisinic acid were found between these A. annua's. For the first time the A. annua plants of different geographical origin were found to belong to different chemotypes. A chemotype with a high artemisinin level was found to have also a high dihydroartemisinic acid level but a relatively low artemisinic acid level. Reversibly, a chemotype with low levels of artemisinin and dihydroartemisinic acid contained a high artemisinic acid level. Artemisinic acid is considered to be the direct precursor of dihydroartemisinic acid in the biosynthetic pathway of artemisinin. The observed accumulation of artemisinic acid in one of the A. annua chemotypes may indicate the presence of a rate-limiting step in the biosynthetic pathway of artemisinin. The enzymatic reduction of artemisinic acid into dihydroartemisinic acid is probably a "bottle neck" in the biosynthetic pathway of artemisinin in varieties with high artemisinic acid and consequentially low artemisinin levels. After a night-frost period, the level of artemisinin was increased, in the Vietnamese A. annua plants, while the dihydroartemisinic acid level was decreased. This phenomenon is in accordance with our hypothesis that stress triggers the conversion of dihydroartemisinic acid to artemisinin. It is suggested that the presence of high levels of dihydroartemisinic acid may be an adaptation to stress conditions (e.g., night-frost), during which relatively high levels of 1O2 are formed. Dihydroartemisinic acid gives the plant protection by reacting with these reactive oxygen species yielding artemisinin as stable end-product.

Water deficit effect on the accumulation of biomass and artemisinin in annual wormwood(Artemisia annua L., Asteraceae)

“…To our knowledge, artemisinin responded to lead and salt-induced oxidative stress by increasing or decreasing according to the doses of the stressors in the soil (Qureshi et al, 2005) or planting in soil with high salinity (Qian et al, 2007); increased in response to a frost (Wallaart et al, 2000); and significantly increased in response to a mild potassium deficiency (Ferreira, 2007). These authors attributed artemisinin accumulation to the generation of active oxygen species in the plant in response to stress, based on the report that high levels of peroxide can be formed in plants under stress triggered by limited water, high UV-B levels, chilling conditions or tissue damage caused by pathogens or frost (Knox and Dodge, 1985).…”

Biotechnology approaches for production of antiulcerogenic dihydro-epideoxyarteannuin B isolated from Artemisia annua L.

“…In the presence of light the compound B was preferentially formed. In accordance to Wallaart et al (2000), artemisinin production occurs in the presence of light continuous by oxidation of the direct precursor dihydroartemisininic hydroperoxide acid and demands a great quantity of 1 O 2 . The authors affi rm that this oxygen form production can take place by some secondary products assigned to chromatophores.…”

“…Heights ( Bowmeester et al (1999) and Wallaart et al (2000), after the conversion of artemisininic acid to dihydroartemisininic acid, all other further conversions up to artemisinin are not enzymatic, but oxidative or photoxidative transformations and thus largely infl uenced by light. The light presence infl uences the terpenes production in both cytoplasmatic and plastidic biosynthetic routes.…”

“…Meanwhile, artemisinin presence has been detected in A. annua transgenic hairy roots in vitro cultivated under light conditions (Liu et al, 2002;Souret et al, 2002;Weathers et al, 2005). Plants can produce high levels of 1 O 2 under stress condition, such as limited water, high UV-B radiation, low temperature, and wounded tissue (Wallaart et al, 2000). Hypothetically, the in vitro culture condition and wounds promoted by cuts in plant tissues could generate a stress condition, exposing root cromatophores to atmospheric air in the fl ask and producing 1 O 2 in the presence of light (Murashige, 1990).…”

“…Wallaart et al (1999) proved that dihydroartemisininic acid and dihydroartemisininic acid hydroperoxide are the direct biosynthetic precursors of artemisinin. Several authors confi rmed that the level of artemisinin and others terpenes production is dependent of factors such as O 2 (Wallaart et al, 2000) and light conditions (Jaziri et al, 1995;Wang et al, 2001;Liu et al, 2002;Weathers et al, 2005). Light can greatly affect the regulation of some genes involved in terpenoids biosynthesis as CLA1 that encodes 1-Deoxy-D-xylulose-5-phosphate syntetase (DXPS) and 1-Deoxy-D-xylulose-5-phosphate reductoisomerase (DXPR).…”