The callus on this medium was whitish green in nature. As NAA at 100mg/l showed maximum induction of andrographolide in leaves, the same concentration of NAA was used to treat the callus of A. paniculata. HPLC analysis of calli treated with NAA showed a peak at 223 nm with the retention time of 4.99 indicating the induction of andrographolide in these samples (Fig.2b). Calli that served as control did not show the peak that represents andrographolide (Fig.2a). This study clearly indicates that andrographolide can be produced under in vitro conditions by treating the callus with NAA.Analogues prepared from natural andrographolide are emerging as powerful anticancer agents. A series of β-amino-γ-butyrolactone analogues synthesized from naturally occurring andrographolide revealed their potential for being developed as promising anti-cancer agents [16]. Further, a novel andrographolide-lipoic acid conjugate (AL-1) was shown to protect pancreatic b-cells from reactive oxygen species (ROS)-induced oxidative injury [17].Present study suggests that andrographolide can be induced in callus and the quantity can be increased by altering the concentration of plant growth hormones used for treating the callus. As callus can be produced throughout the year and is not limited by the time of the year or weather, this approach can be used well in development of andrographolide derivatives against various ailments.
Plant growth hormones play a major role in tissue culture. The composition of nutrient medium along with specific ratio of auxin to cytokinin plays a major role not only in successful establishment of cell cultures but also in the production of secondary metabolites. Rapid high performance thin layer chromatography (HPTLC) screening of callus extracts revealed that the callus established in MS medium supplemented with 4.5 μM NAA and 0.46 μM BAP produced the highest yield of Proscillaridin A (4.51 mg/g DW), Scilliroside (3.3 mg/g DW), Scillaren A (2.35 mg/g DW) and desacetylscilliroside (8.62 mg/g DW), which was higher than from the intact plants of Charybdis congesta[18]. Plant tissue culture systems can be used for continuous production of secondary metabolite compared to whole plants which shows more variation based on physiological and geographical conditions. The quantity of active principle produced can also be increased by changing the concentration of the inducer. Further this approach can considerably decrease the exploitation of medicinal plants where the entire plant is medicinal.