Strategies for indole alkaloids enrichment through callus culture from Alstonia scholaris (L.) R. Br.

The present study was aimed to investigate the effect of different media, plant growth regulators, carbon sources, sucrose concentrations and pH of the medium for indole alkaloids (IAs) accumulation in the callus culture of Alstonia scholaris. The optimized conditions for the accumulation of IAs like echitamine, acetylechitamine, tubotaiwine and picrinine were found to be the MS medium supplemented with 0.3 mg/L of 2,4-D, 0.5 mg/L of FAP and 3% (w/v) sucrose with medium pH 5.8. Efforts were also made to standardize the enrichment protocols of these IAs by modulating concentration and incubation period of various elicitors and precursors. The quantitative analysis of IAs in callus samples was performed by using ultra high-performance liquid chromatography-mass spectrometry. Substantial enrichments of acetylechitamine (6.3780 mg/g DW i.e. ~ 15-fold) and echitamine (1.6513 mg/g DW i.e. ~ 12-fold) were found with 4.5 g/L KCl in 10 days incubation period, followed by tubotaiwine (0.0952 mg/g DW i.e. ~ 4-folds) with 3.0 g/L KCl in 10 days and picrinine (0.3784 mg/g DW i.e. ~ 4-fold) with 4.5 g/L KCl in 2 days.