Optimization of Precursor and Elicitor Utilization in Batch Cultures of Astragalus trojanus Stev.

Öz

Elicitor and precursor applications are commonly used to induce secondary metabolism in plant cell cultures. In this study, methyl jasmonate, jasmonic acid, salicylic acid and pectin were used as elicitors and β-sitosterol was used as a precursor in batch cultures of Astragalus trojanus in order to trigger astragaloside IV and cycloastragenol productions. Growth parameters (fresh and dry weights and dry weight percentages) of batch cultures were also evaluated in order to understand the effects of elicitors and precursor on primary metabolism. All elicitors and precursor used in this study triggered metabolite production at different stages of culture period. The highest astragaloside IV accumulation (0.9435 µg/mg) was detected in medium supplemented with 50 µM methyl jasmonate at the 14th day of culture period, whereas the highest cycloastragenol concentration (0.3626 µg/mg) was found in medium supplemented with 50 µM jasmonic acid at the 28th day of culture period. Large scale cultivation was also performed and 0.3759 µg/mg astragaloside IV was detected in medium supplemented with 50 µM methyl jasmonate at the 14th day.

Anahtar Kelimeler

• Astragalus trojanus,batch culture,elicitor,large-scale production,precursor,primary and secondary metabolism.

Astragalus trojanus Kesikli Kültürlerinde Prekürsör ve Elisitör Kullanımının Optimizasyonu

Öz

Elisitör ve prekürsör uygulamaları bitki hücre kültürlerinde sekonder metabolizmayı hızlandırmak için sıklıkla kullanılan uygulamalardır. Bu çalışmada Astragalus trojanus kesikli kültürlerinde astragalozit IV ve sikloastragenol üretimlerinin arttırılması için metil jasmonat, jasmonik asit, salisilik asit ve pektin elisitör olarak, β-sitosterol ise prekürsör olarak kullanılmıştır. Kesikli kültürlerin büyüme parametreleri (yaş ve kuru ağırlıklar ve kuru ağırlık yüzdeleri) elisitörler ve prekürsörün primer metabolizma üzerine etkilerinin belirlenmesi için değerlendirilmiştir. Elisitörler ve prekürsörün kültür periyodunun farklı basamaklarında metabolit üretimini tetiklediği belirlenmiştir. En yüksek astragalozit IV birikimi (0.9435 µg/mg) 50 µM metil jasmonat içeren besin ortamında kültürün 14. gününde görülürken, en yüksek sikloastragalozit konsantrasyonu 50 µM jasmonik asit içeren besin ortamnda kültürün 28. gününde tespit edilmiştir. Çalışmada büyük ölçekli kültivasyon da gerçekleştirilmiştir ve 50 µM metil jasmonat içeren besin ortamında 14. günde 0.3759 µg/mg astragalozit IV birikimi tespit edilmiştir.

Anahtar Kelimeler

• Astragalus trojanus,kesikli kültür,elisitör,büyük ölçekli üretim,prekürsör,primer ve sekonder metabolizma

Kaynakça

1. Abd El-Mavla, A.M.A. 2010. Production of cycloartane triterpene glycosides in shoot cultures of Astragalus sieberi DC. Nat Prod Res 24(5), 416–422.
2. Angelova, Z., Georgiev, S., Roos, W. 2006. Elicitation of plants. Biotechnol & Biotechnol Eq 20(2), 72-83.
3. Baenas, N., Garcia-Viguera, C., Moreno, D.A. 2014. Biotic elicitors effectively increase the glucosinolates content in Brassicaceae sprouts. J Agric Food Chem 62, 1881−1889.
4. Bedir, E., Çalış, I., Piacente, S., Pizza, C., Khan, I.A. 2000b. New flavonol glycoside from the aerial parts of Astragalus vulneraria. Chem Pharm Bull 48(12), 1994-1995.
5. Bedir, E., Pugh, N., Çalış, I., Pasco, D.S., Khan, I.A. 2000a. Immunostimulatory effects of cycloartane-type triterpene glycosides from Astragalus species. Biol Pharm Bull 23(7), 834-837.
6. Boroujerdnia, M.G., Azemi, M.E., Hemmati, A.A., Taghian, A., Azadmehr, A. 2011. Immunomodulatory effects of Astragalus gypsicolus hydroalcoholic extract in ovalbumin-induced allergic mice model. Iran J Allergy Asthma Immunol 10(4), 281-288.
7. Cai, Y., Luo, Q., Sun, M., Corke, H. 2004. Antioxidant activity and phenolic compounds of 112 traditional Chinese medicinal plants associated with anticancer. Life Sci 74(17), 2157-2184.
8. Cho, J-S., Kim, J-Y., Kim, I-H., Kim, D-I. 2003. Effect of polysaccharide elicitors on the production of decursinol angelate in Agelica gigas Nakai root cultures. Biotechnol & Bioprocess Engineering 8, 158-161.

9. Davis, P.H. 1970. Flora of Turkey and East Aegean Islands, University Press, Edinburgh, Vol. 3, p 164.
10. Du, M., Wu, X.J., Ding, J., Hu, Z.B., White, K.N., Branford-White, C.J. 2003. Astragaloside IV and polysaccharide production by hairy roots of Astragalus membranaceus in bioreactors. Biotechnol Lett 25, 1853-1856.
11. El-Sharabasy, S. 2004. Effects of some precursors on development of secondary products in tissues and media of embryonic callus date palm cv. Sewi. Arab J Biotech 7(1), 83-90.
12. Erişen, S., Atalay, E., Yorgancılar, M. 2011. The effect of thidiazuron on the in vitro shoot development of endemic Astragalus cariensis in Turkey. Turk J Bot 35, 521-526.
13. Erişen, S., Yorgancılar, M., Atalay, E., Babaoğlu, M. 2010. Prolific shoot regeneration of Astragalus cariensis Boiss. Plant Cell Tiss Org Cult 100, 229–233.Hasançebi, S., Turgut-Kara, N., Çakır, Ö., Arı, Ş. 2011. Micropropagation and root culture of Turkish endemic Astragalus chrysochlorus (Leguminosae). Turk J Bot 35, 203–210.
14. Ionkova, I., Kartnig, T., Alfermann, W. 1997. Cycloartane saponin production in hairy root cultures of Astragalus mongholicus. Phytochemistry 45(8), 1597-1600.
15. Jalalpour, Z., Shabani, L., Afghani, L., Sharifi-Tehrani, M., Amini, S-A. 2014. Stimulatory effect of methyl jasmonate and squalestatin on phenolic metabolism through induction of LOX activity in cell suspension culture of yew. Turk J Biol 38, 76-82.
16. Li, W., Fitzloff, J.F. 2001. Determination of astragaloside IV in Radix Astragali (Astragalus membranaceus var. monghulicus) using high-performance liquid chromatography with evaporative light-scattering detection. J Chromatogr Sci 39, 459-462.
17. Lloyd, G.B., McCown, B.H. 1980. Commercial-feasible micropropagation of mountain laurel- Kalmia latifolia by use of shoot- tip culture. Proc Int Plant Prop Soc 30, 421- 427.
18. Misawa, M. 1994. Plant Tissue Culture: An Alternative for Production of Useful Metabolites. FAO Agricultural Services Bulletin No. 108.
19. Nalbantsoy, A., Nesil, T., Erden, S., Çalış, I., Bedir, E. 2011. Adjuvant effects of Astragalus saponins Macrophyllosaponin B and Astragaloside VII. J Ethnopharmacol 134, 897–903.
20. Nalbantsoy, A., Nesil, T., Yılmaz-Dilsiz, Ö., Aksu, G., Khan, S., Bedir, E. 2012. Evaluation of the immunomodulatory properties in mice and in vitro anti-inflammatory activity of cycloartane type saponins from Astragalus species. J Ethnopharmacol 139, 574– 581.
21. Nartop, P., Akay, Ş., Gürel, A. 2013b. Immobilization of Rubia tinctorum L. suspension cultures and its effects on alizarin and purpurin accumulation and biomass production. Plant Cell Tiss Org Cult 112(1), 123-128.
22. Nartop, P., Gurel, A. 2013. Immobilization of Plant Cells and Its Effects on Secondary Metabolite and Biomass Production, Recent Developments in Biotechnology Plant Biotechnology (Multi-Volume Set, Vol.12), Studium Press, ISBN: 0-9656038-5-7.
23. Nartop, P., Gürel, A., Akgün, I.H., Bedir, E. 2013a. Astragaloside IV and cycloastragenol production capacity of Astragalus trojanus calli. Rec Nat Prod 9(1), 49-61.
24. Patel, H., Krishnamurthy, R. 2013. Elicitors in plant tissue culture. J Pharmacognosy & Phytochem 2(2), 60-65.
25. Qian, Z.G., Zhao, Z.J., Xu, Y., Qian, X., Zhong, J.J. 2006. Novel chemically synthesized salicylate derivative as an effective elicitor for inducing the biosynthesis of plant secondary metabolites. Biotechnlog Prog 22(1), 331-3.
26. Savran, T., Gülcemal, D., Masullo, M., Karayıldırım, T., Polat, E., Piacente, S., Alankuş-Çalışkan, Ö. 2012. Cycloartane glycosides from Astragalus erinaceus. Rec Nat Prod 6(3), 230-236.
27. Sevimli-Gür, C., Onbaşılar, I., Atilla, P., Genç, R., Çakar, N., Deliloğlu-Gürhan, I., Bedir, E. 2011. In vitro growth stimulatory and in vivo wound healing studies on cycloartane-type saponins of Astragalus genus. J Ethnopharmacol 134, 844–850.
28. Turgut-Kara, N., Arı, Ş. 2006. Micropropagation of Astragalus maximus Willd. Biotechnol Biotechn Eq 20(1), 20-22.
29. Valenzuela, H.F., Fuller, T., Edwards, J., Finger, D., Molgora, B. 2009. Cycloastragenol extends T cell proliferation by increasing telomerase activity. J Immunol 182, 90–30.
30. Wang, S., Huang, H., Gao, W., Zhuang, C., Li, B., Zhou, P., Kong, D. 2009. Anti-hepatitis B virus activities of astragaloside IV isolated from Radix Astragali. Biol Pharm Bull 32, 132–135.