The study examines the phenolic compounds in hydromethanolic extracts of Salix alba (L.) leaves and bark as well as their antioxidant activity and cytotoxic potential. UPLC-PDA-Q/TOF-MS analysis showed a total of 29 phenolic compounds in leaves and 34 in bark. Total phenolic compound content was 5575.96 mg/100 g of dry weight (DW) in leaves and 2330.31 mg/100 g DW in bark. The compounds were identified as derivatives of phenolic acids (seven in leaves and five in bark), flavanols and procyanidins (eight in leaves and 26 in bark) and flavonols (14 in leaves and three in bark). Both extracts exhibited strong antioxidant potential, assessed by radical scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS), but the bark extract was even stronger than the ascorbic acid used as a standard. The cytotoxicity of both extracts was evaluated against human skin fibroblasts and human epidermal keratinocytes cell lines using the Presto Blue cell viability assay. The keratinocytes were more resistant to tested extracts than fibroblasts. The leaf and bark extracts at concentrations which exhibited antioxidant activity were also not toxic against the keratinocyte cell line. Thus, S. alba extracts, especially the leaf extract, offer promise as a nontoxic natural antioxidant, in cosmetic products or herbal medicines, and as a source of bioactive secondary metabolites.

Rehmannia glutinosa Libosch., a valuable medicinal plant, was successfully propagated in vitro using shoot tip explants. Shoot multiplication was performed in glass tubes and in a nutrient sprinkle bioreactor. A mixture of 0.1 mg L⁻¹ indole-3-acetic acid (IAA) and 1.0 mg L⁻¹ of 6-benzylaminopurine in Murashige and Skoog (MS) agar-solidified medium proved the best combination for multiple shoot induction, yielding 8.2 shoots per explant after 4 weeks of culture in glass tubes. The number of shoots increased to 21 per explant when the same combination of growth regulators was used in a nutrient sprinkle bioreactor. The shoots rooted with a frequency of 93 % after 6 weeks of culture on MS agar medium supplemented with IAA (0.1 mg L⁻¹) before being acclimatized in the greenhouse. The antioxidant activities of methanolic extracts from the leaves and roots of the in vitro-regenerated plants of R. glutinosa cultivated in the greenhouse were evaluated using four in vitro assays: scavenging of free radicals (DPPH and ABTS), transition metal reduction and total antioxidant activity phosphomolybdenum test. In all cases, the methanolic extract from leaves demonstrated better antioxidant activity than those taken from roots. A strong correlation was found between total phenolic and flavonoid content, and the antioxidant capacity of the studied extracts.

The composition of volatile organic compounds emitted by in vitro shoots of Agastache rugosa (Fischer & C.A. Meyer) O. Kuntze (Lamiaceae) was studied using headspace solid-phase microextraction–gas chromatography–mass spectrometry and compared to the those emitted by adult plants and in vitro-germinated seedlings. Shoot-tip explants were cultured on a solid MS medium supplemented with either 4.4 μM 6-benzyladenine (BA), 9.3 μM kinetin, or 0.45 μM thidiazuron and with either 0.57 μM indole-3-acetic acid (IAA) or 0.41 μM picloram. Shoot proliferation was observed in all these treatments. The presence of these plant growth regulators in the culture medium significantly influenced the composition of volatiles as well as morphogenetic responses observed. The number and quality of regenerating shoots and frequency of axillary bud break were highest in medium containing the BA + IAA combination. Sixty-five compounds were identified in the headspace of the in vitro-produced material and plants cultivated in the field. The in vitro shoots emitted both hydrocarbon (limonene, α-pinene) and oxidized (menthone, isomenthone, pulegone) monoterpenes. The composition of monoterpenes differed depending on the type of auxin—rather than cytokinin—in the medium. The emission of phenylallyl compounds, such as estragole, a major compound in field-grown plants, was markedly lower in shoot cultures.

This study is the first description of a procedure for shoot organogenesis from calli derived from various seedling explants. The frequency of shoot induction and number of shoots per explant were measured on Murashige and Skoog (MS) medium with 0.2–3.0 mg L⁻¹ 6-benzylaminopurine (BAP) alone or combined with auxin [0.1 mg L⁻¹ indole-3-acetic acid (IAA) or 1-naphthalenacetic acid]. The highest organogenic response where 90 % of explants formed a regenerative callus with a mean number of nine shoots was achieved after 6 weeks of culture when hypocotyl explants and 1.0 mg L⁻¹ BAP was used together with 0.1 mg L⁻¹ IAA. The shoots regenerated from hypocotyl-derived calli were rooted for 6 weeks on MS medium lacking growth regulators or containing auxin (IAA or indole-3-butyric acid). The plantlets were successfully acclimatized in a greenhouse. Micropropagated plants subjected to random amplified polymorphic DNA and inter simple sequence repeat (ISSR) marker based profiling revealed a uniform banding pattern identical with that of donor plants. The iridoid and phenylethanoid glycoside content of the organogenic callus culture and in vitro-raised plants was determined using UHPLC. In the tested samples catalpol, aucubin, harpagid, harpagoside, verbascoside, isoverbascoside and traces of loganin and catalposide were identified. The leaves produced the highest levels of catalpol (43–45 mg g⁻¹ DW). Organogenic callus and 6-week-old in vitro grown plantlets were characterized by the highest accumulation of aucubin, verbascoside and isoverbascoside. The work also demonstrated the regenerative ability of hypocotyl-derived calli and production of bioactive metabolites to be stable for 4 years of culture.