MICROPROPAGATION STUDIES AND ANTIOXIDANT ANALYSIS OF THE ENDANGERED PLANTS OF BULGARIAN YELLOW GENTIAN (Gentiana lutea L.)

Maria I. Petrova

Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria

Ely G. Zayova

Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria

Lyudmila I. Dimitrova

Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria

Maria P. Geneva

Department of Plant-Soil Interactions, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria

Kamelia D. Miladinova-Georgieva

Department of Plant-Soil Interactions, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria


Abstract

In order to develop an efficient micropropagation system, it is essential to establish the appropriate concentration of growth regulators for seed germination, shoot formation and rooting. Nodal segments from in vitro obtained seedlings of Gentiana lutea L. were cultured in vitro in Murashige and Skoog’s medium supplemented with BAP, Thidiazuron and Zeatin (0.5, 1.0 and 2.0 mg L−1). A maximum number of shoots with the highest height was recorded at 2.0 mg L−1 BAP. For further optimization of the process, we used nutrient media containing BAP and Zeatin with a combination of low concentration of Indoleacetic acid. MS medium containing 2.0 mg L−1 Zeatin and 0.2 mg L−1 IAA resulted in maximum numbers of shoots 94.3) with shoot height 2.5 cm. The multiple plants were successfully ex vitro acclimatized with 65% survival. The presence of growth regulators (2.0 mg L−1 Zeatin and 0.2 mg L−1 IAA) in the nutrient media resulted in an effective antioxidant activity in G. Lutea determined by the low molecular antioxidant metabolites such as phenols and flavonoids and activities of antioxidant enzymes – catalase, ascorbate peroxidase, guaiacol peroxidase, and superoxide dismutase. The described protocol allows the establishment of numerous micropropaged plants of rare and endangered G. lutea.

Keywords:

Gentiana lutea L., seed germination, micropropagation, plant growth regulators, antioxidants

Aberham, A., Schwaiger, S., Stuppner, H., Ganzera, M. (2007). Quantitative analysis of iridoids, secoiridoids, xanthones and xanthone glycosides in Gentiana lutea L. roots by RP-HPLC and LC–MS, J. Pharm. Biomed. Anal., 45, 437–442.

Benson, E., Roubelakis-Angelakis, K. (1992). Fluorescent lipid peroxidation products and antioxidant enzymes in tissue cultures of Vitis vinifera L. Plant Sci., 84, 83–90.

Benzie, I., Strain, J. (1996). The ferric reducing ability of plasma (FRAP) as a measure of “antioxidant power”: The FRAP assay. Anal. Biochem., 239, 70–76.

Bicknell, R. (1984). Seed propagation of Gentiana scabra. Comb. Proc. – Int. Plant Propag. Soc., 34, 396–401.

Bradford, M. (1976). A rapid and sensitive method for the estimation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem., 72, 248–254.

Peev, D. et al. (eds.), (2015). Red Data Book of the Republic of Bulgaria. Vol. 1. Plants and fungi. Bulgarian Academy of Sciences, Ministry of Environment and Waters, Sophia, 1730 pp.

Drobyk, N., Hrytsak, L., Mel’nyk, V., Kravets, N., Konvalyuk, I. Twardovska, M.O., Kunakh, V.A. (2015). In vitro manipulation and propagation of Gentiana L. species from the Ukrainian Flora. In: The Gentianaceae. Vol. 2. Biotechnology and Applications, Rybczynski, J.J., Davey, M.R., Mikula, A. (eds.). Springer, Berlin−Heidelberg, 45–79.

Fiuk, A., Rybczyński, J. (2008). Genotype and plant growth regulator-dependent response of somatic embryogenesis from Gentiana spp. leaf explants. In Vitro Cell. Dev. Biol.–Plant, 44, 90–99.

Gamborg, O., Miller, R., Ojima, K. (1968). Nutrient requirements of suspension cultures of soybean root cells. Exp. Cell Res., 50, 151–158.

Kwak, J., Nguyen, V., Schroeder, J. (2006). The role of reactive oxygen species in hormonal responses. Plant Physiol., 141, 323–329.

Mayorova, O., Hrytsak, L., Drobyk, N. (2015). Adaptation of Gentiana lutea L. plants obtained in vitro to ex vitro and in situ condition. Biotech. Acta, 8, 77–88.

Mirzaee, F., Hosseini, A., Jouybari, H., Davoodi, A., Azadbakht, M. (2017). Medicinal, biological and phytochemical properties of Gentiana species. J. Trad. Compl. Med., 7, 1–9.

Molassiotis, A., Dimassi, K., Diamantidis, G., Therios, I. (2004). Changes in peroxidases and catalase activity during in vitro rooting. Biol. Plant, 48, 1–5.

Momčilović, I., Grubišić, D., Nešković, M. (1997). Micropropagation of four Gentiana species (G. lutea, G. cruciata, G. purpurea and G. acaulis). Plant Cell Tiss. Org. Cult., 49, 141–144.

Morgan, E., Butler, R., Bicknell, R. (1997). In vitro propagation of Gentiana cerina and Gentiana corymbifera. New Zeal. J. Crop Hortic. Sci., 25, 1–8.

Murashige, T., Skoog, F. (1962). A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant, 15, 473–497.

Nastasijevic, B., Lazarevic-Pasti, T., Dimitrijevic-Bran- kovic, S., Pasti, I., Vujacic, A., Joksić, G., Vasić, V. (2012). Inhibition of myeloperoxidase and antioxidative activity of Gentiana lutea extracts. J. Pharm. Biomed. Anal., 66, 191–196.

Niki, E. (2011). Antioxidant capacity: which capacity and how to assess it? J. Berry Res., 1, 169–176.

Ohkawa, K. (1983). Gentiana. In: CRC Hand Book of Flowering, Halevy, A.H. (ed.). CRC Press, Boca Raton, 351–355.

Petrova, M., Zayova, E., Vitkova, A. (2011). Effect of silver nitrate on in vitro root formation of Gentiana lutea. Rom. Biotech. Lett., 16, 53–58.

Pfeffer, H., Dannel, F., Römheld, V. (1998). Are there connection between phenol metabolism, ascorbate metabolism and membrane integrity in leaves of boron-deficient sunflower plants? Physiol. Plant, 104, 479–485.

Šavikin, K., Menkovic´, N., Zdunic´, G., Stevic´, T., Radanovic´, D., Janković, T. (2009). Antimicrobial activity of Gentiana lutea L. extracts. Z. Naturforsch. C, 64, 339–342.

Sidjimova, B., Valyovska-Popova, N., Peev, D. (2014). Reproductive capacity of four medicinal plants in Nature Park “Rilsky Manastir” – West Bulgaria. J. BioSci. Biotechnol., SE/ONLINE, 177–180.

Tepe, B., Sokmen, M., Akpulat, H., Sokmen, A. (2006). Screening of the antioxidant potentials of six Salvia species from Turkey. Food Chem., 95, 200–204.

Tian, M., Gu, Q., Zhu, M. (2003). The involvement of hydrogen peroxide and antioxidant enzymes in the process of shoot organogenesis of strawberry callus. Plant Sci., 165, 701–707.

Viola, U., Franz, C. (1989). In vitro propagation of Gentiana lutea. Planta Med., 55, 690–690.

Zayova, E., Stancheva, I., Geneva, M., Hristozkova, M., Dimitrova, L., Petrova, M., Sichanova, M., Salamon, I., Mudroncekova, S. (2018). Arbuscular mycorrhizal fungi enhance antioxidant capacity of in vitro propagated garden thyme (Thymus vulgaris L.). Symbiosis, 74(3), 177–187.

Zheleznic, A., Baricevic, D., Vodnic, D. (2002). Micropro- pagation and acclimatization of yellow gentian (Gentiana lutea L.). Zb. Biotech. Fac. Univ. Ljubl., 79(1), 253–259.

Zhishen, J., Mengcheng, T., Jianming, W. (1999). The determination of flavonoid contents in mulberry and their scavenging effects on superoxide radicals. Food Chem., 64, 555–559.

Download

Published
2019-06-18



Maria I. Petrova 
Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria
Ely G. Zayova 
Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria
Lyudmila I. Dimitrova 
Department of Applied Genetics and Plant Biotechnology, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria
Maria P. Geneva 
Department of Plant-Soil Interactions, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria
Kamelia D. Miladinova-Georgieva 
Department of Plant-Soil Interactions, Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Acad. George Bonchev, bl. 21, Sofia 1113, Bulgaria



License

 

Articles are made available under the conditions CC BY 4.0 (until 2020 under the conditions CC BY-NC-ND 4.0).
Submission of the paper implies that it has not been published previously, that it is not under consideration for publication elsewhere.

The author signs a statement of the originality of the work, the contribution of individuals, and source of funding.