Anadolu endemik Digitalis L. türlerinin in vitro çoğaltımı ve kardiyak glikozitlerinin üretilmesi

Members of the genus Digitalis L. are medicinally and economically important plants as they contain cardiac glycosides that increase contractions of the heart and regulate heart rhythms. Several methods for high-frequency in vitro regeneration of endemic Digitalis species of Anatolia were established testing different types of basal medium and explants excised from in vitro germinated seedlings. Of those tested medium formulations, Linsmaier and Skoog (1965), supplemented with 0.5 mg/L thidiazuron (TDZ) and 0.25 mg/L indole-3-acetic acid (IAA), was selected mainly for the optimization of later tissue culture protocols of Digitalis davisina Heywood, D. cariensis Jaub. et Spach em. Werner and D. trojana Ivan. For D. lamarckii Ivan., benzyl-amino-purine (BA) and naphtylene-acetic-acid (NAA) were more productive in terms of shoot formation.As an explant source, flamingo-bill type (FBT) explants in general were more productive than other tested hypocotly, leaf or root explants. In addition to shoot regeneration, callus induction was employed in some species (D. cariensis and D. lamarckii) on LS or Murashige and Skoog medium (1962) containing various concentrations of BA and NAA. Callus cultures obtained from D. lamarckii only was able to produce multiple shoots. Regeneration frequency varied from species to species producing a range of 1 to 9 shoots per explant within 6 or 8 weeks of in vitro cultivation periods. Cardenolide profiles of leaf materials of endemic Anatolian Digitalis collected from different locations at various vegetation periods (budding, flowering or seed setting) and of tissue culture samples (leaf, complete shoots, etc.) produced in vitro were also investigated using two instrumental analysis; thin layer chromatography (TLC) and reverse phase high performance liquid chromatography (RP-HPLC). TLC analysis provides us new insights as a fingerprint pattern of the cardenolides depending on various biotic and/or abiotic factors (physiological ages, different locations or vegetation periods). HPLC analysis was substantiated in terms of quantitative analysis of cardenolides detected by TLC. In that, glucogitorosid content, a major cardenolide, found in our endemic species, was directly correlated with plant growth and development by means of exogenously added growth regulators or seasonal factors as well as the geography where they distributed.