Studies on the Production of Digitalis Cardenolides by Plant Tissue Culture HI. Effects of Nutrients on Digitoxin Formation by Shoot-Forming Cultures of Digitalis purpurea L. Grown in Liquid Media

Shoot-forming cultures of Digitalis purpurea L. were grown in various modifications of Murashige-Skoog medium. The effects of vitamins, natural extracts, plant growth substances, carbon and nitrogen sources and phosphate on their growth and digitoxin formation were investigated. Sucrose, glucose and raffinose were suitable carbon sources for both growth and digitoxin formation. The optimum concentration of sucrose was 3%. Reduction of the basal nitrogen concentration by one-third increased the digitoxin content per unit weight without suppression of growth. The optimal ratio of nitrate-nitrogen to ammonium-nitrogen was 2, as provided in the basal medium. A threefold increase in the phosphate concentration improved both growth and digitoxin content. ThiamineHCl and myo-inositol were required for digitoxin formation, whereas glycine, nicotinic acid and pyridoxineHCl, usual components of the medium, were not required for either growth or digitoxin formation during the three passages examined. Coconut milk improved growth with no reduction in the digitoxin content per unit weight. Gibberellic and abscisic acids at 0.01 to 0.1 mg per liter slightly improved digitoxin formation. Kinetin had no clear positive effect on either growth or digitoxin formation