Elicitation and permeabilisation affect the accumulation and storage profile of phytoestrogens in high productive suspension cultures of Genista tinctoria

Abstract

High productive suspension cultures of Genista tinctoria were elicited (methyl jasmonate or chitosan) and permeabilised (dimethyl sulfoxide) in order to achieve a plant in vitro system rich in isoflavones, with extracellular storage profile. The maximum concentration of isoflavone aglycones (4-6 times higher than in the controlled biomass) was obtained in the suspension elicited with chitosan. All isoflavonoids were stored inside cells. In case of methyl jasmonate supplementation the total concentration of isoflavone aglycones achieved was the result of the de novo biosynthesis as well as a hydrolysis of the storage ester forms. The presence of chitosan in the medium was only associated with the production of aglycones de novo. The elicitors had no effect on the accumulation and metabolism of the basic glycoside isoflavones in the suspension. The change in the way the isoflavones were stored was achieved after supplementing the growth media with dimethyl sulfoxide. Maximum concentration of isoflavones in the medium was observed in the 7th hour of the experiment. Eventually, a plant growth system was developed, producing over 11% of isoflavones, ejected as a result of chemical permeabilisation, into the growth medium (approx. 80% of total amount of these compounds in the biomass).

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

“…However, the result has implications for studies aimed at establishing large-scale systems for the production of isoflavones. As described in recent work by the authors (Luczkiewicz and Kokotkiewicz 2012), G. tinctoria callus grown on the above medium proved suitable for establishing high-productive suspension cultures (isoflavone content up to ca. 11 % dry weight), which can be further used for bioreactor studies.…”

“…With isoflavone content exceeding 10 % dry weight, callus cultures of this species maintained on SH medium supplemented with 0.5 mg l -1 TIBA and 5.0 mg l -1 KIN are so far the richest source of isoflavonoids among in vitro cultures of higher plants. In the previous research (Luczkiewicz and Kokotkiewicz 2012), the authors have already succeeded in establishing high-productive suspension cultures of G. tinctoria using liquid medium with the same PGR composition. However, further scale-up studies in bioreactors are required to establish an efficient system for isoflavone production.…”

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

“…Even though these constituents were not found in C. subternata suspension, it produced a set of isoflavone glucosides (CG, PG and FG), which were formerly found in callus of this species but not in the intact plant material (Kokotkiewicz et al 2012b). No isoflavones were detected in the growth medium, reflecting the typical, intracellular storage profile of the above compounds (Goyal and Ramawat 2008a, b; Sharma et al 2009; Shinde et al 2009a; Luczkiewicz and Kokotkiewicz 2012). …”

“…(Li and Zhang 2006; Goyal and Ramawat 2008a, b; Sharma et al 2009), Psoralea sp. (Shinde et al 2009a), Glycine max (Federici et al 2003; Terrier et al 2007) and Genista tinctoria (Luczkiewicz and Glod 2005; Luczkiewicz and Kokotkiewicz 2012). These are characterised mainly by the presence of genistein, genistin, daidzein and puerarin, which belong to the best known phytoestrogens (Patisaul and Jefferson 2010).…”

Improved production of dibenzocyclooctadiene lignans in the elicited microshoot cultures of Schisandra chinensis (Chinese magnolia vine)

“…In the course of our experiments on S. chinensis microshoots, the biomass was elicited with MeJa at 50, 100, and 200 μM on the 23rd and/or the 27th day of the growth cycle and collected on the 30th day (Figure S3 , Table S2 ). Moreover, the experiments included cadmium chloride (CdCl 2 ) (Figure S4 , Table S3 ), which was previously shown to increase the accumulation of biologically active lignans (phyllanthin and hypophyllanthin) in Phyllanthus amarus (Rai et al 2005 ), and dimethylsulfoxide (DMSO) (Figure S2 , Table S1 ), demonstrated to act as both elicitor of secondary metabolism (Mannan et al 2010 ) and an effective permeabilizing agent (Luczkiewicz and Kokotkiewicz 2012 ; Jaremicz et al 2014 ). CdCl 2 was applied at 2.5, 5, 10, and 20 mM, and DMSO was added at 0.2, 2, 4, and 8% v/v .…”