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Resumen de Biotechnological production of rosmarinic acid and ruscogenins. Scaling up the optimized processes to benchtop bioreactors

Abbas Khojasteh

  • Rosmarinic acid (RA), an ester of caffeic acid and 3,4-dihydroxyphenyl lactic acid, is widely distributed in the plant kingdom, including Satureja khuzistanica, a threatened Iranian plant species. Interest in RA is growing due to its biological activities, including cognitive-enhancing effects, slowing the development of Alzheimer’s disease, cancer chemoprotection and anti-inflammatory activity. In order to meet the increasing demand for this compound and preserve its natural sources, a new biotechnological platform based on cell suspension cultures of S. khuzistanica was developed. The results show that cell suspensions of this plant species synthesized high amounts of RA, which accumulated mainly inside the cells. In order to improve the system’s biotechnological production, the effect of three elicitors, methyl jasmonate (MeJA), cyclodextrin (CD) and coronatine (COR) were tested. The obtained results demonstrate the effectiveness of MeJA and COR in improving the RA production of the S. khuzistanica cell cultures and also that the time course of growth and RA production can change with the age of the cell line and the number of subcultures. In contrast, the effects of CD were found to be insignificant. In this pipeline, the scale up of the biotechnological system to a benchtop bioreactor demonstrated the adaptability of the S. khuzistanica cell cultures to the scaling-up process and that the elicitors MeJA and COR also effectively improved the production at bioreactor level. Concerning the biological activity of S. khuzistanica, we demonstrated that an enriched RA extract was able to reduce the viability of cancer cell lines, increasing the sub-G0/G1 cell population and the activity of caspase-8 in MCF-7 cells, which suggests that S. khuzistanica extracts can induce apoptosis of MCF-7 cells through activation of the extrinsic pathway. In addition, our findings indicate that other compounds of the extract may act synergistically to potentiate the anticancer activity of RA.

    Similarly to S. khuzistanica, Ruscus aculeatus is a threatened medicinal plant. Its main bioactive components, the steroid saponins ruscogenin and neoruscogenin, have long been used in the treatment of hemorrhoids and varicose veins, but recently demonstrated activity against some types of cancer have stimulated interest in R.

    aculeatus extracts. Thus, plant cell biofactories could constitute an alternative to the whole plant as a source of these bioactive compounds. In our work, despite the in vitro recalcitrance of R. aculeatus, after many attempts in which we tested more than 70 different culture media, we developed friable R. aculeatus calli and their derived plant cell suspensions. Their ruscognenin production was compared with that of organized in vitro plantlet and root-rhizome cultures. Root-rhizomes showed a higher capacity for biomass and ruscogenin production than the cell suspensions and the yields were greatly improved by elicitation with COR, as in other plant in vitro cultures. The final total ruscogenin content was higher than 1800 mg g DW-1, after 4 weeks of culture. Although ruscogenins accumulate in plants mainly in the root-rhizome, it was demonstrated that the aerial part could play an important role in their biosynthesis, because production was higher in the whole plant than in the root-rhizome cultures. The most productive biotechnological system, COR-elicited root rhizome cultures, was scaled up to a benchtop bioreactor without losing its capacity for biomass and ruscogenin production.


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