Micropropagation and HPLC-DAD, UPLC MS/MS analysis of oenothein B and phenolic acids in shoot cultures and in regenerate
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ORIGINAL ARTICLE
Micropropagation and HPLC-DAD, UPLC MS/MS analysis of oenothein B and phenolic acids in shoot cultures and in regenerated plants of fireweed (Chamerion angustifolium (L.) Holub) Mariola Dreger1 · Agnieszka Gryszczyńska2 · Milena Szalata1 · Karolina Wielgus1 Received: 4 May 2020 / Accepted: 12 October 2020 © The Author(s) 2020
Abstract In this study, a micropropagation protocol using nodal explants from in vitro grown plants of Chamerion angustifolium (L.) Holub was developed and analysis of oenothein B and selected phenolic acids in shoot cultures was performed for the first time. For shoot induction and multiplication Murashige and Skoog’s (MS) basal medium supplemented with 2-isopentenyladenine (2iP), zeatin (Z) and 6-benzyloaminopurine (BAP) was used. 2iP was the most responsive in terms of promoting shoots per explant with the maximum (6.57 ± 1.14) recorded at a concentration of 2.0 mg L −1 after 6 weeks of culture. After two subcultures the multiplication rate was increased up to 19 shoots per explant on medium with 2iP (1.0 mg L−1). To prevent tissue browning, ascorbic acid and casein hydrolysate were added to the induction medium, resulting in a reduction of browning by 30%. The rooted plantlets were successfully transferred to soil and acclimatized with 97% frequency. Quantitative and qualitative assessments of oenothein B and phenolic acid contents in in vitro regenerated shoots as well as in ex vitro plants were performed using high-performance liquid chromatography with a diode-array detector (HPLC-DAD) and ultra-performance liquid chromatography–tandem mass spectrometry (UPLC MS/MS) methods. Oenothein B (1.62‒4.55 g 100 g−1 DW), ellagic acid, gallic and caffeic acids were identified in in vitro regenerated plants. The results of this study confirm that the oenothein B-producing plantlets can be obtained using the micropropagation method with axillary shoots being a valuable source of oenothein B and phenolic acids. Key message An efficient regeneration protocol from nodal explants of fireweed (Chamerion angustifolium (L.) Holub) was developed for the first time. The presence of oenothein B and phenolic acids in regenerated shoots and ex vitro cultivated plants was confirmed using HPLC-DAD and UPLC analysis. Keywords Chamerion angustifolium (L.) Holub · Nodal explants · Shoot cultures · HPLC-DAD · UPLC MS/MS · Oenothein B
Introduction Communicated by Ali R. Alan. * Mariola Dreger [email protected] 1
Department of Biotechnology, Institute of Natural Fibres and Medicinal Plants, Wojska Polskiego 71B, 60‑630 Poznań, Poland
Department of Pharmacology and Phytochemistry, Institute of Natural Fibres and Medicinal Plants, Kolejowa 2, 62‑064 Plewiska, Poland
2
Fireweed (Chamerion angustifolium (L.) Holub, Chamaenerion angustifolium (L.) Scop. syn. Epilobium angustifolium L.) is an herbaceous perennial plant from the Onagraceae family. The genus Chamerion is a separate monophyletic group with eight species restricted to the northern hemisphere (Wagner et al. 2007). Fire
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