Regulation of vincamine biosynthesis and associated growth promoting effects through abiotic elicitation, cyclooxygenase inhibition, and precursor feeding of bioreactor grown Vinca minor hairy roots.

Abstract

Hydroxylase/acetyltransferase elicitors and cyclooxygenase inhibitor along with various precursors from primary shikimate and secoiridoid pools have been fortified to vincamine less hairy root clone of Vinca minor to determine the regulatory factors associated with vincamine biosynthesis. Growth kinetic studies revealed that acetyltransferase elicitor acetic anhydride and terpenoid precursor loganin significantly reduce the growth either supplemented alone or in combination (GI = 140.6 ± 18.5 to 246.7 ± 24.3), while shikimate and tryptophan trigger biomass accumulation (GI = 440.2 ± 31.5 to 540.5 ± 40.3). Loganin also downregulates total alkaloid biosynthesis. Maximum flux towards vincamine production (0.017 ± 0.001% dry wt.) was obtained when 20-day-old hairy roots were fortified with secologanin (10mg/l) along with tryptophan (100mg/l), naproxen (8.4mg/l), hydrogen peroxide (20μg/l), and acetic anhydride (32.4mg/l). This was supported by RT PCR (qPCR) analysis where 2- and 3-fold increase in tryptophan decarboxylase (TDC; RQ = 2.0 ± 0.09) and strictosidine synthase (STR; RQ = 3.3 ± 0.36) activity, respectively, was recorded. The analysis of variance (ANOVA) for growth kinetics, total alkaloid content, and gene expression studies favored highly significant data (P < 0.05-0.01). Above treated hairy roots were also up-scaled in a 5-l stirred-tank bioreactor where a 40-day cycle yielded 8-fold increase in fresh root mass.