Isoflavone production in Cyclopia subternata Vogel (honeybush) suspension cultures grown in shake flasks and stirred-tank bioreactor

Głód

2014

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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, b-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 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.

Section: Discussionmentioning

confidence: 94%

Plant growth regulators affect biosynthesis and accumulation profile of isoflavone phytoestrogens in high-productive in vitro cultures of Genista tinctoria

Głód

2014

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“…Chinese plants of the genus Astragalus, considered a major source of free and glucosidated calycosin, are threatened by overexploitation because of slow growth of the roots combined with high market demand for natural medicines (Wu et al 2011;Xu et al 2011). Cell cultures of Cyclopia subternata were previously shown to accumulate 7-O-b-glucosides of calycosin, pseudobaptigenin and formononetin, absent in intact plant material (Kokotkiewicz et al 2012(Kokotkiewicz et al , 2013, and can thus be utilized for the production of these derivatives independently of their natural resources. In the present work, the effect of varying light and temperature regimes on biomass growth and accumulation of bioflavonoids in C. subternata callus cultures (Kokotkiewicz et al 2009) was investigated.…”

mentioning

confidence: 99%

“…1). The samples were freeze-dried, extracted and assessed for the production of isoflavonoids (CG, PG, FG) and flavonoids (H), identified as major phenolic metabolites in the investigated cultures (Kokotkiewicz et al 2009(Kokotkiewicz et al , 2012), using the previously described HPLC method (Kokotkiewicz et al 2009(Kokotkiewicz et al , 2013.…”

mentioning

confidence: 99%

Light and temperature conditions affect bioflavonoid accumulation in callus cultures of Cyclopia subternata Vogel (honeybush)

Buciński

2014

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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-b-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.

“…These cultures are deposited in the Higher Plants Biotechnology Laboratory at the Department of Pharmacognosy, Medical University of Gdansk, Poland, and is available to other researchers. As demonstrated in previous study (Kokotkiewicz et al 2012), C. genistoides microshoots accumulated xanthones (M and IM), flavanone hesperidin (H), as well as three isoflavone glucosides [7-O-b-glucosides of calycosin (CG), formononetin (CG) and pseudobaptigenin (PG)], absent in intact plant material but otherwise characteristic for undifferentiated cultures of Cyclopia (Kokotkiewicz et al 2013a(Kokotkiewicz et al , 2014. However, since the current research focused on establishing an in vitro source of xanthones and benzophenones, the experiments were conducted using SH medium enriched with indole-3-butyric acid (IBA), which was previously shown to enhance the accumulation of phenolics typical for field-grown plants (Kokotkiewicz et al 2012).…”

mentioning

confidence: 97%

“…The length of growth period (60 d) was set based on the results of the previous studies concerning C. genistoides micropropagation which showed substantial increase in xanthone levels in explants elongated for 2 months on IBA-supplemented medium (Kokotkiewicz et al 2012). The harvested shoots were evaluated for polyphenol content according to the previously described methodology (Kokotkiewicz et al 2012(Kokotkiewicz et al , 2013a.…”

mentioning

confidence: 99%

Xanthone, benzophenone and bioflavonoid accumulation in Cyclopia genistoides (L.) Vent. (honeybush) shoot cultures grown on membrane rafts and in a temporary immersion system

Buciński

2014

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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-b-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.