Pyruvate and α-ketoglutarate precursor feeding combined with methyl jasmonate elicitation increases anthraquinone accumulation in hairy root cultures of Rubia tinctorum

Anthraquinones (AQs) are specialized metabolites produced by Rubia tinctorum that have traditionally been used as dyes, but also have important biological activities. The present research studied the effects of the addition of pyruvate (Pyr) and α-ketoglutarate (KG) precursors in combination with methyl jasmonate (MeJa) elicitation, via a full factorial design, on hairy root cultures of R. tinctorum . MeJa was the main factor that triggered AQ accumulation, and the addition of Pyr and KG also significantly enhanced AQ production. Feeding of each precursor resulted in increased intracellular and total AQ production, which revealed that Pyr and KG are limiting precursors of AQ biosynthesis in hairy root cultures of R. tinctorum . The three variables showed a significant interaction effect, resulting in total AQ production of ~ 2022 µM. Moreover, the addition of Pyr and KG increased AQ release into the culture medium. KG was the main factor responsible for the secretion of AQs; combination with MeJa elicitation resulted in 56 µM of AQs in the culture medium. The phenolic and flavonoid contents were also evaluated, and the results showed that MeJa elicitation triggered only flavonoid production. At the same time, precursor addition had no significant effect on phenolic or flavonoid accumulation.