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

Applied Microbiology and Biotechnology, Nov 2017

Dibenzocyclooctadiene lignans are a specific group of secondary metabolites that occur solely in Schisandra chinensis. The aim of the presented work was to boost the accumulation of lignans in the agitated microshoot cultures of S. chinensis, using different elicitation schemes. The experiments included testing of various concentrations and supplementation times of cadmium chloride (CdCl2), chitosan (Ch), yeast extract (YeE), methyl jasmonate (MeJa), and permeabilizing agent—dimethylsulfoxide (DMSO). After 30 days, the microshoots were harvested and evaluated for growth parameters and lignan content by LC-DAD method. The analyses showed enhanced production of lignans in the elicited S. chinensis microshoots, whereas the respective media samples contained only trace amounts of the examined compounds (< 5 mg/l). Elicitation with CdCl2 caused up to 2-fold increase in the total lignan content (max. ca. 730 mg/100 g DW after the addition of 1000 μM CdCl2 on the tenth day). Experiments with chitosan resulted in up to 1.35-fold increase in lignan concentration (max. ca. 500 mg/100 g DW) after the supplementation with 50 mg/l on the first day and 200 mg/l on the tenth day. High improvement of lignan production was also recorded after YeE elicitation. After the elicitation with 5000 mg/l of YeE on the first day of the growth period, and with 1000 and 3000 mg/l on the 20th day, the lignan production increased to the same degree—about 1.8-fold. The supplementation with 1000 mg/l YeE on the 20th day of the growth cycle was chosen as the optimal elicitation scheme, for the microshoot cultures maintained in Plantform temporary immersion system—the total content of the estimated lignans was equal to 831.6 mg/100 g DW.

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

Improved production of dibenzocyclooctadiene lignans in the elicited microshoot cultures of Schisandra chinensis (Chinese magnolia vine) Agnieszka Szopa 0 1 Halina Ekiert 0 1 Adam Kokotkiewicz 0 1 Agata Król 0 1 Maria Luczkiewicz 0 1 0 Department of Pharmacognosy, Faculty of Pharmacy, Medical University of Gdańsk , al. gen. J. Hallera 107, 80-416 Gdańsk , Poland 1 Department of Pharmaceutical Botany, Jagiellonian University , Collegium Medicum, ul. Medyczna 9, 30-688 Kraków , Poland Dibenzocyclooctadiene lignans are a specific group of secondary metabolites that occur solely in Schisandra chinensis. The aim of the presented work was to boost the accumulation of lignans in the agitated microshoot cultures of S. chinensis, using different elicitation schemes. The experiments included testing of various concentrations and supplementation times of cadmium chloride (CdCl2), chitosan (Ch), yeast extract (YeE), methyl jasmonate (MeJa), and permeabilizing agent-dimethylsulfoxide (DMSO). After 30 days, the microshoots were harvested and evaluated for growth parameters and lignan content by LC-DAD method. The analyses showed enhanced production of lignans in the elicited S. chinensis microshoots, whereas the respective media samples contained only trace amounts of the examined compounds (< 5 mg/l). Elicitation with CdCl2 caused up to 2-fold increase in the total lignan content (max. ca. 730 mg/100 g DW after the addition of 1000 μM CdCl2 on the tenth day). Experiments with chitosan resulted in up to 1.35-fold increase in lignan concentration (max. ca. 500 mg/100 g DW) after the supplementation with 50 mg/l on the first day and 200 mg/l on the tenth day. High improvement of lignan production was also recorded after YeE elicitation. After the elicitation with 5000 mg/l of YeE on the first day of the growth period, and with 1000 and 3000 mg/l on the 20th day, the lignan production increased to the same degree-about 1.8-fold. The supplementation with 1000 mg/l YeE on the 20th day of the growth cycle was chosen as the optimal elicitation scheme, for the microshoot cultures maintained in Plantform temporary immersion system-the total content of the estimated lignans was equal to 831.6 mg/100 g DW. Elicitation; Temporary-immersion bioreactor; Schisandra lignan production; Biotic elicitors; Abiotic elicitors; In vitro cultures Introduction Schisandra chinensis (Turcz.) Baill., Chinese magnolia vine (Schisandraceae), is a climbing plant, naturally occurring in the countries of Eastern Asia, whereas in European countries, it grows mainly as an ornamental shrub (Panossian and Wikman 2008; Szopa et al. 2016a; Szopa et al. 2017a) . The raw material of Schisandra fruits, Schisandrae chinensis fructus (chin. běi wǔ wèi zi; literally Bfive-flavor berry^), has been used for therapeutic purposes in traditional Chinese medicine and has successfully been included in pharmacopoeial monographs of Asian and European countries as well as in the USA Pharmacopoeia and the International Pharmacopoeia, printed by WHO (World Health Organization 2007; European Directorate for the Quality of Medicines. 2017; Szopa et al. 2017a) . Numerous therapeutic properties of Schisandra fruit extracts, as well as its individual compounds, have been confirmed by the scientific research, carried out with the use of in vivo and in vitro models. The studies indicate that S. chinensis has a positive effect on liver functioning and stimulates cardiovascular, respiratory, and central nervous systems. The therapeutic properties of Chinese magnolia vine include anticancer, immunostimulant, and adaptogenic (Mocan et al. 2016; Szopa et al. 2016a; Szopa et al. 2017a) . These activities are related to the presence of the dibenzocyclooctadiene lignans, known as BSchisandra lignans^ (Figure S1), due to the fact that their occurrence is limited to S. chinensis (Fuss 2004; Opletal et al. 2004; Lu and Chen 2009) . The dominant lignans that can be distinguished out of over 40 present in S. chinensis are as follows: schisandrin; gomisins A, C, F and N; deoxyschisandrin; and γ-schisandrin. The extracts of S. chinensis fruit have been used for the manufacture of medical products, food supplements, or cosmetics (Szopa et al. 2016a) . Given the fact that the lignans of S. chinensis are therapeutically unique compounds, the research on alternative methods of their natural resource-independent acquisition has been performed. The methods of the chemical synthesis of Schisandra lignans have been developed; however, these had limited success due to their complex stereochemistry (Shi et al. 2009) . The other alternative is plant cell culture which has so far been employed for the production of several biologically active secondary metabolites, including lignans. Moreover, it has been reported that in vitro techniques enable to enhance the production of lignans by means of elicitation, addition of biosynthetic precursors, and immobilization (Angelova et al. 2006; Ver (...truncated)


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Agnieszka Szopa, Adam Kokotkiewicz, Agata Król, Maria Luczkiewicz, Halina Ekiert. Improved production of dibenzocyclooctadiene lignans in the elicited microshoot cultures of Schisandra chinensis (Chinese magnolia vine), Applied Microbiology and Biotechnology, 2017, pp. 1-15, DOI: 10.1007/s00253-017-8640-7