In vitro regeneration and cardenolide determination of an endemic foxglove, Digitalis cariensis (Aegean Foxglove)

The genus Digitalis L. is one of the most pharmacologically important plant genera because many Digitalis species produce cardenolides, which are commonly used for cardiac insufficiencies. In this study, different pretreatments were employed to increase germination of Digitalis cariensis seeds, which germinate poorly because of extensive dormancy. Seed germination was improved when seeds were pretreated by scarification followed by soaking in sterile distilled water overnight. In addition, the regeneration capacity of four different types of explants (cotyledonary leaf, root, hypocotyl, or flamingo-bill-type [FBT]) was tested on Linsmaier and Skoog (LS), Murashige and Skoog (MS) and Gamborg's B5 (B5) medium containing no plant growth regulators or supplemented with 0.5 mg L⁻¹ thidiazuron (TDZ) alone or in combination with 0.25 mg L⁻¹ indole-3-acetic acid (IAA). Among the explants tested, only FBT explants regenerated. The highest mean number of shoots was produced on LS medium supplemented with a combination of 0.5 mg L⁻¹ TDZ and 0.25 mg L⁻¹ IAA, producing 3.9 shoots per FBT explant after 4 wk of incubation. Regenerated shoots were rooted on LS medium containing different concentrations of IAA or indole-3-butyric acid (IBA) (0.1, 0.5, or 1.0 mg L⁻¹). The greatest number of roots developed on medium supplemented with 1.0 mg L⁻¹ IAA, which produced 5.6 roots per shoot after 3 wk of culture. Cardenolides were profiled by HPLC analysis from basal leaves of D. cariensis plants from natural populations and from in vitro-derived plantlets. There was no significant difference in lanatoside contents (A, B, and C) between the two sources. Digoxin and digitoxin were not detected in either source.