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. 2018 Jul 3;23(7):1619.
doi: 10.3390/molecules23071619.

Isoflavones Production and Possible Mechanism of Their Exudation in Genista tinctoria L. Suspension Culture after Treatment with Vanadium Compounds

Milan Skalicky  1 , Jan Kubes  2   3 , Vaclav Hejnak  4 , Lenka Tumova  5 , Jaroslava Martinkova  6 , Jan Martin  7 , Helena Hnilickova  8
Affiliations

Isoflavones Production and Possible Mechanism of Their Exudation in Genista tinctoria L. Suspension Culture after Treatment with Vanadium Compounds

Milan Skalicky et al. Molecules. .

Abstract

The family Fabaceae traditionally serves as a food and herbal remedies source. Certain plants serve for treatment of menopausal symptoms based on a presence of typical secondary metabolites, isoflavones. Beside soybean and clovers, other plants or cultures in vitro can produce these molecules. A cultivation in vitro can be enhanced by elicitation that stimulates metabolites biosynthesis via stress reaction. Vanadium compounds have been already described as potential elicitors, and the aim of this study was to determine the impact of NH₄VO₃ and VOSO₄ solutions on isoflavones production in Genista tinctoria L. cell cultures. The significant increase of isoflavones content, such as genistin, genistein, or formononetin, was measured in a nutrient medium or dry mass after NH₄VO₃ treatment for 24 or 48 h. The possible transport mechanism of isoflavones release as a result of elicitation was further evaluated. An incubation with different transport inhibitors prior to elicitation took effect on isoflavones content in the medium. However, there was a non-ended result for particular metabolites such as genistein and daidzein, where ATP-binding cassette (ABC) or, alternatively, multidrug and toxin extrusion (MATE) proteins can participate. Possible elicitation by some inhibitors was discussed as a result of their pleiotropic effect. Despite this outcome, the determination of the transport mechanism is an important step for identification of the specific transporter.

Keywords: Dyer’s Greenweed; elicitation; heavy metals; plasma membrane transport.

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Conflict of interest statement

The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; and in the decision to publish the results.

Figures

Figure 1
Figure 1
Effects of elicitors on isoflavone content in nutrient medium (NM) and dry matter (DM) after 24 or 48 h. Genista tinctoria cell culture was treated by 1 μM (a) or 10 μM (b) of NH4VO3 and 1 μM (c) or 10 μM (d) of VOSO4. Bars indicate isoflavone levels in treated samples recalculated to a relative 100% isoflavone content in water control samples. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between tested and water control samples within Tukey’s test.
Figure 2
Figure 2
Effects of NH4Cl on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 3
Figure 3
Effects of gramicidin on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 4
Figure 4
Effects of brefeldin A on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 5
Figure 5
Effects of Na3VO4 on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 6
Figure 6
Effects of verapamil on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE.
Figure 7
Figure 7
Effects of probenecid on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 8
Figure 8
Effects of glibenclamide on isoflavone content in nutrient medium (NM) and dry matter (DM) of Genista tinctoria cell culture. Zero concentration of inhibitor represents water control. All bar values were recalculated to a relative 100% isoflavone content in samples with NH4VO3 (1 μM) after 24 h. Data are mean of three repeats ± SE; (*) represent significant difference (p < 0.05) between elicited and inhibited or water control samples within Tukey’s test.
Figure 9
Figure 9
HPLC chromatogram of monitored isoflavones standards and their chemical formula.
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