Effect of growth regulators on transformed root cultures of Hyoscyamus muticus

The transformed root cultures of Hyoscyamus muticus are usually cultivated in hormone-free medium in which the production of hyoscyamine is high. We studied whether the addition of exogenous growth regulators could enhance tropane alkaloid accumulation without affecting the growth and root phenotype. In order to rapidly screen a large number of hairy root samples a monoclonal antibody based enzyme immunological assay was developed for scopolamine analysis. The hairy root cultures tolerated exogenous auxins in the concentration range 0.01-5 mu mol/L well. The growth of the roots was not significantly affected by IAA and NAA, but the alkaloid accumulation was doubled compared with that of the roots growing in the absence of auxin. At optimum IAA concentration (0.1-1.0 mu mol/L) the maximum hyoscyamine content was 117 mg L-1 and scopolamine content 0.2 mg L-1. In general, the scopolamine contents were by a factor of 300-500 lower than the hyoscyamine ones. The phenotype, growth rate and alkaloid accumulation of the roots did not change despite cytokinin addition. Both ABA and GA(3) strongly inhibited hyoscyamine accumulation. However, the inhibition was not associated with the changes in root morphology. We suggest that endogenous auxin production in the roots is not sufficient for maximum tropane alkaloid accumulation. To our knowledge this is the first time that alkaloid accumulation has been significantly increased by exogenous auxins without influencing growth.