Genetic transformation of Datura stramonium for the production of tropane alkaloids in hairy root cultures

Abstract

Datura stramonium is well known for its tropane alkaloid content, specifically atropine and scopolamine. With the demand of tropane alkaloids in the pharmaceutical sector, as anti cholinergic agents, anesthetic or its use in transdermal patches, an alternate method to obtain higher yields is required. In addition, much interest has been developed around the plant for its potential biological profile due to its use in traditional medicine. This study investigated the potential production of tropane alkaloids in plant tissue cultures and the biological activity in crude extracts of D. stramonium. Elicited hairy root cultures had displayed the highest atropine and scopolamine content as compared to in vitro leaf and callus culture extracts. The concentrations of atropine and scopolamine was 5.2 µg.mL-1 and 5.01 µg.mL-1 respectively. This was an increase of 1.57 and 1.21 fold respectively as compared to wild plant extracts. The biological profile of D. stramonium was investigated by determining its antimicrobial activity and anti-oxidant activity. The anti bacterial activity wsas investigated by the Kirby-Bauer disc diffusion assay. Root extracts displayed a broader spectrum of bactericidal activity (7 mm – 33 mm) as compared to leaf extracts. No fungicidal activity was displayed by both extracts. The anti oxidant activity was investigated using 2-diphenyl-1-picrylhydrazyl (DPPH) photometric assay. The activity displayed was dose dependant, i.e.; as the extract concentration increased, so did the anti oxidant activity. At a concentration of 1 µg.mL-1 , the radical scavenging capacity of root and leaf extracts was 64.4% and 31.7% respectively whereas, at the highest concentration of 1000 µg.mL-1 , the radical scavenging capacity of root extracts and leaf extracts was 98.4% and 45.8% respectively. The biosafety evaluation of leaf and root extract demonstrated a linear relationship between the concentration of extracts and percentage toxicity. As the concentration and exposure period of the extracts increased, the toxicity towards A. salina had also increased. These results indicated the potential of plant, cell and tissue culture systems of D. stramonium to produce higher yields of atropine and scopolamine through optimization, and the potential of anti oxidant and anti bacterial components present in its leaf and root.