Abstract
The work aimed on studying the effect of synthetic phytohormones on growth and biosynthetic parameters of a suspension culture of Tribulus terrestris L., and the influence of cytokinins on these characteristics. In addition to the previously obtained results about alternative action of two types of auxins, 2,4-D and alpha-NAA (the first of them activates cell growth in vitro but inhibits the formation of steroidal glycosides, while the second causes cytodifferentiation, leading to a slowdown and arrest of culture growth, but activates the formation of secondary compounds), it was shown that the type of cytokinin and its concentration can modulate its action. It was found that, after transferring a culture from a "growth" medium (containing 2,4-D) to a medium with alpha-NAA and a different composition of cytokinins (BAP or kinetin in various concentrations), it retains the ability to actively grow for at least three growing cycles. Then, with a suboptimal combination of phytohormones, the cell culture stops growing and dies. However, with a certain composition of growth regulators in a nutrient medium (1 or 2 mg/L alpha-NAA + 1 mg/L kinetin), after a long lag phase (12-18 days), culture growth resumes and its long-term cultivation is subsequently possible. Suspension on a medium with 2 mg/L alpha-NAA and 1 mg/L kinetin T. terrestris were maintained for 70 growing cycles without deterioration of growth characteristics (the maximum level of accumulation of dry biomass M-max was 12-16 g/L, growth index I was 13-18, specific growth rate mu was 0.18-0.28 days(-1)). In this case, the content of steroidal glycosides in cells did not fall below 0.2-0.3% of dry biomass. Using the UPLC-ESI-MS method, the presence in the cell biomass of at least four steroidal glycosides of the furostanol type-derivatives of hydroxy-diosgenin and tigogenin/neotigogenin-was shown. The results obtained confirm the general patterns of development of cell populations in vitro and are important for optimizing the growth and biosynthetic characteristics of plant cell cultures.
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