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Microshoots of the East Asian medicinal plant species Schisandra chinensis (Chinese magnolia vine) were grown in bioreactors characterized by different construction and cultivation mode. The tested systems included two continuous immersion systems—a cone-type bioreactor (CNB) and a cylindric tube bioreactor (CTB), a nutrient sprinkle bioreactor (NSB), and two temporary immersion systems (TIS)—RITA® and Plantform. Microshoots were grown for 30 and 60 days in the MS medium enriched with 1 mg l−1 NAA and 3 mg l−1 BA. The accumulation of two groups of phenolic compounds: phenolic acids and flavonoids in the bioreactor-grown S. chinensis biomass, was evaluated for the first time. In the microshoot extracts, seven phenolic acids: chlorogenic, gallic, p–hydroxybenzoic, protocatechuic, syringic, salicylic and vanillic, and three flavonoids: kaempferol, quercitrin and rutoside, were identified. The highest total amount of phenolic acids (46.68 mg 100 g−1 DW) was recorded in the biomass maintained in the CNB for 30 days. The highest total content of flavonoids (29.02 mg 100 g−1 DW) was found in the microshoots maintained in the NSB for 30 days. The predominant metabolites in all the tested systems were: gallic acid (up to 10.01 mg 100 g−1 DW), protocatechuic acid (maximal concentration 16.30 mg 100 g−1 DW), and quercitrin (highest content 21.00 mg 100 g−1 DW). The influence of bioreactor type on the accumulation of phenolic acids and flavonoids in microshoot cultures of Schisandra chinensis was proven and optimized.

In the presented work, plant growth regulators (PGRs) were tested for their influence on biomass growth and accumulation of therapeutically-relevant indolizidine alkaloids in Securinega suffruticosa (Phyllanthaceae) callus cultures. The study included 9 auxins [2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid, indole-3-acetic acid (IAA), 4-chlorophenoxyacetic acid, indole-3-butyric acid, 4-amino-3,5,6-trichloropicolinic acid, β-naphthoxyacetic acid, 1-naphthaleneacetic acid, indole-3-propionic acid, one polar auxin transport inhibitor (2,3,5-triiodobenzoic acid) and 7 cytokinins [2-isopentenyladenine, 1-phenyl-3-(1,2,3-thiadiazol-5-yl)urea, N-(2-chloro-4-pyridinyl)-N′-phenylurea, diphenylurea, zeatin, kinetin (KIN), 6-benzylaminopurine]. The respective PGRs were applied at 0.5 and 5.0 mg l−1, together with 5.0 or 0.5 mg l−1 KIN or 2,4-D (for auxins and cytokinins, respectively). The calli subjected to different phytohormone combinations (39 modifications in total) were evaluated for growth and alkaloid content, and the results tested for statistical differences. The highest concentrations of securinine (1.73 mg g−1 DW) and allosecurinine (3.11 mg g−1 DW) were recorded in the callus grown in the presence of 0.5 mg L−1 IAA and 5.0 mg L−1 KIN. To the best of the authors’ knowledge, this is the first such comprehensive report concerning the influence of PGRs on alkaloid accumulation in plant in vitro cultures.

The influence of plant growth regulators on biomass growth and the accumulation of medicinally-relevant isoflavone phytoestrogens, derivatives of genistein and daidzein (8 compounds including aglycones, glucosides and glucoside esters) in callus cultures of Genista tinctoria (Fabaceae) was examined. The experiments included 10 auxins [2,4-dichlorophenoxyacetic acid (2,4-D), p-chlorophenoxyacetic acid, indole-3-acetic acid, indole-3-butyric acid, indole-3-propionic acid, 1-naphthaleneacetic acid, β-naphthoxyacetic acid, picloram, 2,3,5-triiodobenzoic acid (TIBA), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T)] and 7 cytokinins [6-benzylaminopurine, forchlorfenuron, 1,3-diphenylurea, 2-isopentenyladenine, kinetin (KIN), thidiazuron, zeatin] applied at 0.5 and 5.0 mg l−1, jointly with 5.0 or 0.5 mg l−1 KIN or 2,4-D (for auxins and cytokinins, respectively—36 phytohormone combinations in total). Statistical analysis of the relationships between callus growth [expressed as growth index (Gi)] and the accumulation of isoflavones showed positive correlation in the cytokinin group (rxy values from 0.13 to 0.61) and negative correlation within auxins (rxy values from −0.31 to −0.39). Among the cytokinins tested, the highest isoflavone content (6,436.26 mg/100 g dry weight) and the fastest biomass growth (Gi = 892.46 %) were obtained for 0.5 mg l−1 KIN used jointly with 5.0 mg l−1 2,4-D. In the group of auxins, the combination of 0.5 mg l−1 TIBA and 5.0 mg l−1 KIN provided the fastest culture growth (Gi = 983.07 %) and the isoflavone concentration of 10,474.23 mg/100 g dry weight, which is so far the highest amount of these metabolites achieved in callus cultures of higher plants.

In vitro shoots of the commercially important South-African legume Cyclopia genistoides were grown in different liquid culture systems [membrane rafts (MR) and a temporary immersion bioreactor (TIB)] and evaluated for the accumulation of phenolic secondary metabolites. The major constituents of the investigated cultures were medicinally relevant xanthones [mangiferin (M) and isomangiferin (IM)] and benzophenone derivatives [iriflophenone 3-C-β-glucoside (IG)]. The highest concentrations of M, IM and IG in MR-grown shoots were 1,843.59, 712.02 and 594.29 mg 100 g−1 dry wt, respectively. Bioreactor cultivation provided higher peak concentrations of M (2,622.70 mg 100 g−1 dry wt), IM (757.40 mg 100 g−1 dry wt) and IG (648.30 mg 100 g−1 dry wt) which corresponded to the respective productivities of 5.48, 1.58 and 3.04 mg l−1 d−1. The results indicate that TIB cultures of C. genistoides may be utilized as an alternative source of the above constituents, particularly IM and IG, which are relatively expensive and so far hardly available from commercial sources.

Callus cultures of the endemic South-African legume Cyclopia subternata were cultivated under varying light and temperature conditions to determine their influence on biomass growth and bioflavonoids accumulation. Experimental modifications of light included complete darkness, light of different spectral quality (white, red, blue and yellow) and ultraviolet C (UVC) irradiation. The calli were also subjected to elevated temperature or cold stress. Among the tested light regimes, cultivation under blue light resulted in the highest levels of hesperidin (H)—118.00 mg 100 g−1 dry weight (DW) on 28 days of experiment, as well as isoflavones: 7-O-β-glucosides of calycosin (CG), pseudobaptigenin (PG) and formononetin (FG)—28.74, 19.26 and 10.32 mg 100 g−1 DW, respectively, in 14-days old calli. UVC irradiation applied on 20 days stimulated the accumulation of H (204.14 mg 100 g−1 DW), CG (31.84 mg 100 g−1 DW) and PG (18.09 mg 100 g−1 DW) in 28 days culture by 140, 46 and 165 %, respectively, without negatively influencing callus growth. Low temperature (13 °C) increased CG content by over 1,500 % (235.29 mg 100 g−1 DW) when applied during the whole 28-days growth cycle, at the same time causing 95 % decrease in culture growth in comparison to reference calli maintained at 24 °C. On the contrary, elevated temperature (29 °C) applied during the second half of the culture period resulted in over 300 and 500 % increase in CG and PG content (61.76 and 58.89 mg 100 g−1, respectively) while maintaining relatively high biomass yield.