Establishment of In Vitro Cell Suspension Culture, Kinetics of Cell Growth, pH, Nutrient Uptake and Production of 2-hydroxy-4-Methoxybenzaldehyde from the Germinated Root of Decalepis hamiltonii Wight & Arn. -An Endangered Plant

Abstract

Decalepis hamiltonii Wight & Arn. belongs to Asclepiadaceae family which is an ethno-pharmaceutically important monogenic plant species that is native to the Deccan peninsula forest areas of India. It is endangered due to habitat loss and over exploitation for volatile 2-hydroxy-4-methoxybenzaldehyde (HMB), which is an aromatic bioactive secondary metabolite. HMB is of great biological significance and is present in the plant’s tuberous roots. Plant cell culture is a viable alternative method for in vivo plant cultivation for secondary metabolites production. Callus induction with high biomass from the germinated root as explants for HMB production was optimised on Murashige and Skoog’s (MS) medium containing 3% (w/v) sucrose, supplemented with 1 μM 2,4-dichlorophenoxyacetic acid (2,4-D), 1 μM α- naphthaleneacetic acid (NAA) and 10 μM 6-benzylaminopurine (BAP) in dark incubation. Cell suspension cultures were established in 250 ml shake flasks, and each flask contained 50 ml of the same induction medium. Moreover, the pH 5.8, 3% sugar concentration and 120 rpm agitation speed in dark condition were suitable for high biomass production and the specific growth rate (µ) was 0.086 /day. Extraction of HMB was done by steam condensate methods from the biomass of cell suspension culture. Qualitative and quantification analyses of HMB were performed with gas chromatography (GC) and observed that 0.92 ± 0.02 mg/ml (0.092%) HMB were synthesized in the cell suspension culture.