Agitated shoot cultures of three Hypericum perforatum cultivars: 'Elixir', 'Helos' and 'Topas' were established and maintained on Linsmaier and Skoog (LS) and Murashige and Skoog (MS) media containing varying concentrations (0.1-3.0 mg l -1 ) of a-naphthaleneacetic acid (NAA) and 6-benzyladenine (BA). In methanolic extracts of the biomass, the amounts of free phenolic acids and cinnamic acid were determined by HPLC. Six of the seventeen compounds analyzed were detected in all the extracts: 3,4-dihydroxyphenylacetic acid, p-coumaric acid, protocatechuic acid, syringic acid, and two depsideschlorogenic and neochlorogenic acids. The amounts of individual compounds and the total amount of phenolic acids depended on the media variants used and the increases ranged from 1.2-to 15.0-fold and from 1.2-to 1.9-fold, respectively. The maximum total amounts in the biomass of 'Elixir', 'Helos' and 'Topas' were 212, 222, and 191 DW). The richest source of the estimated compounds and hence a potential biotechnological platform were the shoots of 'Helos' cultivated on LS and MS media containing low concentrations of NAA and BA (0.1-1.0 mg l -1 ).

Methanolic extracts from in vitro grown Scutellaria lateriflora shoots cultured on five Murashige and Skoog (MS) medium variants supplemented with different combinations of 6-benzylaminopurine (BAP) and α-naphthaleneacetic acid (NAA) under different light conditions (monochromatic light, white light and no light) were analysed by HPLC for three groups of metabolites: flavonoids (26 compounds), phenolic acids and their precursors (19+2) and phenylethanoid glycosides (2). The analyses revealed the presence of baicalein, baicalin, wogonin, wogonoside, 3,4-dihydroxyphenylacetic acid and verbascoside. There was clear evidence of the influence of plant growth regulators and light conditions on the accumulation of the analysed groups of secondary metabolites. The amounts of the compounds changed within a wide range—for the total flavonoid content, 30.2-fold (max. 1204.3 mg·100 g−1 dry weight (DW)); for 3,4-dihydroxyphenylacetic acid, 5.5-fold (max. 33.56 mg·100 g−1 DW); and for verbascoside, 1.5-fold (169.15 max. mg·100 g−1 DW). The best medium for the production of most of the compounds was the Murashige and Skoog variant with 1 mg l−1 BAP and 1 mg l−1 NAA. For verbascoside, the best ‘productive’ medium was the MS variant supplemented with 0.5 mg l−1 BAP and 2 mg l−1 NAA. The accumulation of the metabolites was stimulated to the greatest extent by blue light, under which the extracts were found to contain the highest total amount of flavonoids and the highest amounts of flavonoid glucuronides, baicalin and wogonoside, as well as of verbascoside. Their amounts were, respectively, 1.54-, 1.49-, 2.05- and 1.86-fold higher than under the control white light.

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Methanolic extracts of microshoots from agar cultures and of microshoots and media from agitated cultures of Scutellaria lateriflora grown on identical five variants of the Murashige-Skoog (MS) and Linsmaier-Skoog (LS) media with different 6-benzyladenine (BA) and 1-naphthaleneacetic acid (NAA) concentrations (0.5-3.0 mg/l) were analyzed for flavonoids (27 compounds), phenylpropanoid glycosides (2 compounds) and phenolic acids (19 compounds) using the DAD-HPLC method. The amounts of individual compounds and total amounts of the above mentioned metabolite groups in the biomass from both types of the tested cultures were dependent on the composition of basal media and concentrations of BA and NAA in the media variants. In media extracts, no metabolites were confirmed. Higher total amounts of flavonoids were obtained in agar cultures (max. 722.04 and 2989.55 mg/100 g DW on MS and LS media variants, respectively), which were 1.1 and 1.7 times higher, respectively, than in agitated cultures. In both types of cultures, glycosidic conjugates: baicalin and wogonoside were the main flavonoids (max. amounts: 513.93 and 1838.18 mg/100 g DW, and 305.92 and 700.85 mg/100 g DW in agar cultures on MS and LS variants, respectively). High amounts of verbascoside in agar cultures were also confirmed (max. 384.99 and 543.17 mg/100 g DW on MS and LS media variants, respectively). The cells grown in agitated cultures converted exogenous precursors, i.e. hydroquinone (HQ) and 4-hydroxybenzoic acid (4-HBAc) into arbutin (β-d-glucoside of HQ). The highest amount of this product reached 5.63 and 1.45 g/100 g DW after the addition of these two precursors, respectively. This is the first large-scale report documenting in vitro biosynthetic potential of Scutellaria lateriflora microshoots cultivated in two tested culture. Key message After testing of 10 media variants in two types of in vitro cultures high amounts of flavonoids (app. 3 g%) and verbascoside (0.54 g%) were obtained. Additionally, 5.6 g% of arbutin as hydroquinone β-d-glucosylation product was confirmed. Keywords American skullcap • In vitro culture • Scutellaria specific flavonoids • Verbascoside • Arbutin • HPLC analysis Abbreviations BA 6-Benzyladenine 3,4-DAc 3,4-Dihydroxyphenylacetic acid DAD-HPLC Diode-array detector-high performance liquid chromatography DW Dry weight FM Fresh biomass 4-HBAc 4-Hydroxybenzoic acid HQ Hydroquinone LS Linsmaier and Skoog MS Murashige and Skoog NAA 1-Naphthaleneacetic acid Communicated by Nokwanda Pearl Makunga.

In this study we assess the effect of pyrrolizidine alkaloids (PAs) extracted from Lithospermum canescens on the biology of the two-spotted spider mite (Tetranychus urticae Koch). Lithospermum canenscens (Michaux) Lehm. (Boraginaceae) is a common prairie plant also known as Indian paint or hoary puccoon. A mixture of seven PAs with known chemical structures was used in this investigation. Mites treated with PAs showed a high mortality of juveniles, a decrease in female fecundity and a shortened longevity. The intrinsic rate of population increase (r(m)) was used as an indicator of T. urticae population performance after treatment with PAs. The r(m) value obtained with alkaloid-treated leaves was lower than that for mites developing on untreated leaves, which indicates that the mite population would develop much slower on treated plants. The results suggest that further studies should be performed to assess the possible use of PA extracts for spider mite control.