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Production Enhancement of Medicinally Active Coumarin and Phenolic Compounds in Hairy Root Cultures of Pelargonium sidoides: The Effect of Elicitation and Sucrose

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Abstract

African geranium (Pelargonium sidoides DC., Geraniaceae) is a herbaceous perennial plant with well-known medicinal properties. In the present study, the effect of elicitation by methyl jasmonate (MeJ) (25, 50, and 100 µM) and sucrose (15 and 30 g/L) on the fresh weight (FW), dry weight (DW) and the production of pharmaceutically active coumarin (umckalin) and phenolics (gallic, vanillic, and p-coumaric acids) in P. sidoides hairy root cultures was investigated. Hairy roots were induced from the leaves of in vitro sterile P. sidoides seedlings inoculated with Agrobacterium rhizogenes strains (LBA-9402, C58C1) and Agrobacterium tumefaciens C58C1 (pRiA4). Among the fifteen evaluated hairy root lines (HRLs), HRL4 showed the highest accumulation of biomass (9.6 g FW and 0.56 DW) and was selected for elicitation and sucrose management for 50 days in a shaking system. The treated HRs, developed in Murashige and Skoog liquid media, demonstrated the highest FW (32.68 g) and DW (1.1 g) 50 days after elicitation with 100 µM of MeJ. However, a 1.4-fold enhancement of gallic and vanillic acids was achieved with 50 and 25 µM of MeJ, respectively, in HRs under control conditions. The highest accumulation of umckalin (1.61-fold increase compared to the control) was obtained with 100 µM of MeJ. These findings can be useful to develop commercial production of umckalin and phenolics in P. sidoides HR cultures in scaled-up systems and bioreactors.

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Abbreviations

DPE:

Days post elicitation

FW:

Fresh weight

DW:

Dry weight

HRCs:

Hairy root cultures

HRLs:

Hairy root lines

HPLC:

High-performance liquid chromatography

MeJ:

Methyl jasmonate

SMs:

Secondary metabolites

MS:

Murashige and Skoog

PCR:

Polymerase chain reaction

References

  • Aftab T, Khan MMA, Idrees M, Naeem M, Moinuddin HN (2011) Methyl jasmonate counteracts boron toxicity by preventing oxidative stress and regulating antioxidant enzyme activities and artemisinin biosynthesis in Artemisia annua L. Protoplasma 248:601–612

    CAS  PubMed  Google Scholar 

  • Ansari MA, Chung I, Rajakumar G, Alzohairy MA, Almatroudi A, Khanna V, Thiruvengadam M (2019) Evaluation of polyphenolic compounds and pharmacological activities in hairy root cultures of Ligularia fischeri Turcz. f. spiciformis (Nakai). Molecules 24(8):1586

    CAS  Google Scholar 

  • Baskan S, Oztekin N, Bedia Erim F (2007) Determination of carnosic acid and rosmarinic acid in Sage by capillary electrophoresis. Food Chem 101:1748–1752

    CAS  Google Scholar 

  • Brendler T, Van Wyk BE (2008) A historical, scientific and commercial perspective on the medicinal use of Pelargonium sidoides (Geraniaceae). J Ethnopharmacol 119:420–433

    CAS  PubMed  Google Scholar 

  • Bladt S, Wagner H (2007) From Zulu medicine to the European phytomedicine Umckaloabo®. Phytomedicine 14(1):2–4

    PubMed  Google Scholar 

  • Bulgakov VP, Khodakovskaya MV, Labetskaya NV, Chernoded GK, Zhuravlev YN (1998) The impact of plant rolC oncogene on ginsenoside production by ginseng hairy root cultures. Photochem 49:1929–1934

    CAS  Google Scholar 

  • Bulgakov VP, Tchernoded GK, Mischenko NP, Khodakovskaya MV, Glazunov VP, Zvereva EV, Fedoreyev SA, Zhuravlev YN (2002) Effects of salicylic acid, methyl jasmonate, etephone and cantharidin on anthraquinone production by Rubia cordifolia callus cultures transformed with rol B and rol C genes. J Biotechnol 97:213–221

    CAS  PubMed  Google Scholar 

  • Careddu D, Pettenazzo A (2018) Pelargonium sidoides extract EPs 7630: a review of its clinical efficacy and safety for treating acute respiratory tract infections in children. Int J Gen Med 11:91–98

    CAS  PubMed  PubMed Central  Google Scholar 

  • Casanova E, Trillas MI, Moysset L, Vainstein A (2005) Influence of rol genes in floriculture. Biotechnol Adv 23:3–39

    CAS  PubMed  Google Scholar 

  • Chandra S, Chandra R (2011) Engineering secondary metabolite production in hairy roots. Phytochem Rev 10(3):371

    CAS  Google Scholar 

  • Cheruvathur MK, Thomas TD (2014) Effect of plant growth regulators and elicitors on rhinacanthin accumulation in hairy root cultures of Rhinacanthus nasutus (L.) Kurz. Plant Cell Tiss Org Cult 118:169–177

    CAS  Google Scholar 

  • Colling J, Groenewald JH, Makunga NP (2010) Genetic alterations for increased courmarin production lead to metabolic changes in the medicinally important Pelargonium sidoides DC. (Geraniaceae). Metab Eng 12:561–572

    CAS  PubMed  Google Scholar 

  • Chung I, Rekha K, Rajakumar G, Thiruvengadam M (2016a) Production o glucosinolates, phenolic compound and associated gene expression profiles of hairy root culture in turnip (Brassica rapa ssp. Rapa). 3 Biotech 6:175

    PubMed  PubMed Central  Google Scholar 

  • Conceicao LFR, Ferreres F, Tavares RM, Dias ACP (2006) Induction of phenolic compounds in Hypericum perforatum L. cells by Colletotrichum gloeosporioides elicitation. Phytochemistry 67:149–155

    CAS  PubMed  Google Scholar 

  • Chung IM, Thiruvengadam M, Rekha K, Rajakumar G (2016b) Elicitation enhanced the production of phenolic compounds and biological activities in hairy root cultures of bitter melon (Momordica charantia L.). Braz Arch Biol Technol 59:1–10

    Google Scholar 

  • De Wet H, Van Heerden FR, Van Wyk BE (2005) Alkaloids of Antizoma miersiana (Menispermaceae). Biochem Syst Ecol 33:799–807

    Google Scholar 

  • Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem Bull 19:11–15

    Google Scholar 

  • Duchow S, Blaschek W, Classen B (2015) Reproduction of the medicinal plant Pelargonium sidoides via somatic embryogenesis. Planta Med 81(12/13):1169–1174

    CAS  PubMed  Google Scholar 

  • Franco L, de Oliveira BH (2010) Determination of umckalin in commercial tincture and phytopreparations containing Pelargonium sidoides by HPLC: comparison of sample preparation procedures. Talanta 81(4–5):1368–1372

    CAS  PubMed  Google Scholar 

  • Frankfater CHR, Dowd MK, Triplett BA (2009) Effect of elicitors on the production of gossypol and methylated gossypol in cotton hairy roots. Plant Cell Tiss Org Cult 98:341–349

    CAS  Google Scholar 

  • Gabr A, Mabrok H, Abdel-Rahim E, ElBahr M, Smetanska I (2018) Determination of lignans, phenolic acids and antioxidant capacity in transformed hairy root culture of Linum usitatissimum. Nat Prod Res 15:1867–1871

    Google Scholar 

  • Georgiev MI, Pavlov AI, Bley T (2007) Hairy root type plant in vitro systems as sources of bioactive substances. Appl Microbiol Biotechnol 74:1175–1185

    CAS  PubMed  Google Scholar 

  • Grzegorczyk-Karolak I, Kuźma Ł, Skała E, Kiss AK (2018) Hairy root cultures of Salvia viridis L. for production of polyphenolic compounds. Ind Crops Prod 117:235–244

    CAS  Google Scholar 

  • Guillon S, Tre´mouillaux-Guiller J, Pati PK, Rideau M, Gantet P, (2006) Hairy root research: recent scenario and exciting prospects. Curr Opin Plant Biol 9:341–346

    CAS  PubMed  Google Scholar 

  • Helfer M, Koppensteiner H, Schneider M, Rebensburg S, Forcisi S, Muller C, Schmitt-Kopplin Ph, Schindler M, Brack-Werner R (2014) The root extract of the medicinal plant Pelargonium sidoides is a potent HIV-1 attachment inhibitor. PLoS ONE 9(1):e87487

    PubMed  PubMed Central  Google Scholar 

  • Jiao J, Gai QY, Wang X, Qin QP, Wang ZY, Liu J, Fu YJ (2018) Chitosan elicitation of Isatis tinctoria L. hairy root cultures for enhancing flavonoid productivity and gene expression and related antioxidant activity. Ind Crop Prod 124:28–35

    CAS  Google Scholar 

  • Kim OT, Bang KH, Shin YS, Lee MJ, Jung SJ, Hyun DY, Kim YC, Seong NS, Cha SW, Hwang B (2007) Enhanced production of asiaticoside from hairy root cultures of Centella asiatica (L.) Urban elicited by methyl jasmonate. Plant Cell Rep 26:1941–1949

    CAS  PubMed  Google Scholar 

  • Kim YK, Li X, Xu H, Park NI, Uddin MR, Pyon JY, Park SU (2009) Production of phenolic compounds in hairy root culture of tartary buckwheat (Fagopyrum tataricum Gaertn). J Crop Sci Biotechnol 12:53–58

    Google Scholar 

  • Kolodziej H, Kiderlen AF (2007) In vitro evaluation of antibacterial and immunomodulatory activities of Pelargonium reniforme, Pelargonium sidoides and the related herbal drug preparation EPss 7630. Phytomedicine 14:18–26

    CAS  PubMed  Google Scholar 

  • Kostova I, Momekov G, Tzanova T, Karaivanova M (2006) Synthesis, Characterization, and cytotoxic activity of new lanthanum(III) complexes of bis-coumarins. Bioinorg Chem Appl

  • Lewu FB, Adebola PO, Afolayan AJ (2007) Commercial harvesting of Pelargonium sidoides in the Eastern Cape, South Africa: striking a balance between resource conservation and rural livelihoods. J Arid Environ 70:380–388

    Google Scholar 

  • Mayet M (2010) Bio piracy under fire: the Pelargonium patent hearing: a briefing paper by the African center for biosafety /www.biosafetyafrica.netS. Accessed 20 July 2010.

  • Memelink J (2009) Regulation of gene expression by jasmonate hormones. Photochemistry 70:1560–1570

    CAS  Google Scholar 

  • Moyo M, Van Staden J (2014) Medicinal properties and conservation of pelargonium sidoides DC. J Ethnopharmacol 152:243–255

    PubMed  PubMed Central  Google Scholar 

  • Mikkelsen MD, Petersen BL, Glawischnig E, Jensen AB, Andreasson E, Halkier BA (2003) Modulation of CYP79 genes and glucosinolate profiles in Arabidopsis by defense signaling pathways. Plant Physiol 131:298–308

    CAS  PubMed  PubMed Central  Google Scholar 

  • Mirjalili HM, Fakhr Tabatabaei SM, Bonfill M, Alizadeh H, Cusido RM, Ghassempour A, Palazon J (2009) Morphology and withanolide production of Withania coagulans hairy root cultures. Eng Life Sci 9(3):197–204

    CAS  Google Scholar 

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

    CAS  Google Scholar 

  • Narayani M, Srivastava S (2017) Elicitation: a stimulation of stress in in vitro plant cell/ tissue cultures for enhancement of secondary metabolite production. Phytochem Rev 16:1227–1252

    CAS  Google Scholar 

  • Neto VBP, Otoni WC (2003) Carbon sources and their osmotic potential in plant tissue culture: does it matter? Sci Hortic 97:193–202

    Google Scholar 

  • Newton D, Raimondo D, Motjotji L, Lippai C (2013) Biodiversity management plan for Pelargonium sidoides DC. Notice 433 of 2013. Department of Environ- mental Affairs. Republic of South Africa.

  • Ono NN, Tian L (2011) The multiplicity of hairy root cultures: prolific possibilities. Plant Sci 180:439–446

    CAS  PubMed  Google Scholar 

  • Palazón J, Cusidó RM, Gonzalo J, Bonfill M, Morales S, Pinol MT (1998) Relation between the amount the rolC gene product and indole alkaloid accumulation in Catharantus roseus transformed root cultures. J Plant Physiol 153:712–718

    Google Scholar 

  • Palazón J, Mallol A, Eibl R, Lettenbauer C, Cusido RM, Pinol MT (2003) Growth and ginsenside production in hairy root cultures of Panax ginseng using a novel bioreactor. Planta Med 69:344–349

    PubMed  Google Scholar 

  • Pellegrineschi A, Damon JP, Valtorta N, Paillard N, Tepfer D (1994) Improvement of ornamental characters and fragrance production in lemonscented geranium through genetic transformation by Agrobacterium rhizogenes. Biotechnology 12:64–68

    CAS  Google Scholar 

  • Piątczak E, Jeleń A, Makowczyńska J, Zielińska S, Kuźma Ł, Balcerczak E (2019) Establishment of hairy root cultures of Rehmannia elata NE Brown ex Prain and production of iridoid and phenylethanoid glycosides. Ind Crops Prod 137:308–314

    Google Scholar 

  • Praveen N, Murthy HN (2012) Synthesis of withanolide A depends on carbon source and medium pH in hairy root cultures of Withania somnifera. Ind Crop Prod 35:241–243

    CAS  Google Scholar 

  • Ramirez-Estrada K, Vidal-Limon H, Hidalgo D, Moyano E, Golenioswki M, Cusidó RM, Palazon J (2016) Elicitation, an effective strategy for the biotechnological production of bioactive high-added value compounds in plant cell factories. Molecules 21(2):182

    PubMed  PubMed Central  Google Scholar 

  • Satdive RK, Fulzele DP, Eapen S (2007) Enhanced production of azadirachtin by hairy root cultures of Azadirachta indica A. Juss by elicitation and media optimization. J Biotechnol 128:281–289

    CAS  PubMed  Google Scholar 

  • Schulz V (2008) Dose finding study on the efficacy of Pelargonium sidoides extract (EPs® 7630 in new tablet form) in 405 adults with acute bronchitis. Z Phytother 29:37–38

    Google Scholar 

  • Shukla AK, Shasany AK, Verma RK, Gupta MM, Mathur AK, Khanuja SPS (2010) Influence of cellular differentiation and elicitation on intermediate and late steps of terpenoid indole alkaloid biosynthesis in Catharanthus roseus. Protoplasma 242:35–47

    CAS  PubMed  Google Scholar 

  • Sirvent T, Gibson D (2002) Induction of hypericins and hyperforin in Hypericum perforatum L. in response to biotic and chemical elicitors. Physiol Mol Plant Pathol 60:311–320

    CAS  Google Scholar 

  • Srivastava S, Srivastava AK (2007) Hairy root culture for mass-production of high-value secondary metabolites. Crit Rev Biotechnol 27:29–43

    CAS  PubMed  Google Scholar 

  • Srivastava M, Singh G, Sharma S, Shukla S, Misra P (2019) Elicitation enhanced the yield of glycyrrhizin and antioxidant activities in hairy root cultures of Glycyrrhiza glabra L. J Plant Growth Regul 38(2):373–384

    CAS  PubMed  Google Scholar 

  • Thwe A, Valan Arasu M, Li X, Park CH, Kim SJ, Al-Dhabi NA, Park SU (2016) Effect of different Agrobacterium rhizogenes strains on hairy root induction and phenylpropanoid biosynthesis in tartary buckwheat (Fagopyrum tataricum Gaertn). Front Microbial 7:318

    Google Scholar 

  • Thiruvengadam M, Praveen N, John KM, Yang YS, Kim SH, Chung IM (2014) Establishment of Momordica charantia hairy root cultures for the production of phenolic compounds and determination of their biological activities. Plant Cell Tiss Org Cult 118:545–557

    CAS  Google Scholar 

  • Thiruvengadam M, Kim SH, Chung IM (2015) Exogenous phytohormones increase the accumulation of health-promoting metabolites, and influence the expression patterns of biosynthesis related genes and biological activity in Chinese cabbage (Brassica rapa spp. pekinensis). Sci Hortic 193:136–146

    CAS  Google Scholar 

  • Thiruvengadam M, Baskar V, Kim SH, Chung IM (2016) Effects of abscisic acid, jasmonic acid and salicylic acid on the content of phytochemicals and their gene expression profiles and biological activity in turnip (Brassica rapa ssp. rapa). Plant Growth Regul 80:377–390

    CAS  Google Scholar 

  • Udomsuk L, Jarukamjorn K, Tanaka H, Putalun W (2009) Isoflavonoid production in a hairy roots culture of Pueraria candollei. Z Naturforsch C 64:687–691

    CAS  PubMed  Google Scholar 

  • Verpoorte R, Contin A, Memelink J (2002) Biotechnology for the production of plant secondary metabolites. Phytochem Rev 1:13–25

    CAS  Google Scholar 

  • Verma PC, Singh H, Negi AS, Saxena G, Rahman LU, Banerjee S (2015) Yield enhancement strategies for the production of picroliv from hairy root culture of Picrorhiza kurroa Royle ex Benth. Plant Signal Behav 10(5):e1023976

    PubMed  PubMed Central  Google Scholar 

  • White AG, Davies-Coleman MT, Ripley BS (2008) Measuring and optimizing umckalin concentration in wild-harvested and cultivated Pelargonium sidoides (Geraniaceae). S Afr J Bot 74(2):260–267

    CAS  Google Scholar 

  • Weremczuk-Jezyna I, Grzegorczyk-Karolak I, Frydrych B, Krolicka A, Wysokinska H (2013) Hairy roots of Dracocephalum moldavica: rosmarinic acid content and antioxidant potential. Acta Physiol Plant 35:2095–2103

    CAS  Google Scholar 

  • Yousefian Z, Hosseini B, Rezadoost H, Palazon J, Mirjalili MH (2018) Production of the anticancer compound withaferin A from genetically transformed hairy root cultures of withania somnifera. J Nat Prod Commun 13:943–948

    Google Scholar 

  • Zaker A, Sykora C, Gössnitzer F, Abrishamchi P, Asili J, Mousavi SH, Wawrosch C (2015) Effects of some elicitors on tanshinone production in adventitious root cultures of Perovskia abrotanoides Karel. Ind Crop Prod 67:97–102

    CAS  Google Scholar 

  • Zdravkovic-Korac S, Muhovski Y, Druartm P, Calic D, Radojevic L (2004) Agrobacterium rhizogenes-mediated DNA transfer to Aesculus hippocastanum L. and the regeneration of transformed plants. Plant Cell Rep 22:698–704

    CAS  PubMed  Google Scholar 

  • Zhang H, Memelink J (2009) Regulation of secondary metabolism by jasmonate hormones. In Plant-derived natural products (pp. 181–194). Springer, New York. https://doi.org/10.1007/978-0-387-85498-4-8.

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Acknowledgements

This project was financially supported by Iran National Science Foundation (INSF) for financial support of this project with the Grant Number #96003588. The authors thanks Pars Gita Daru Company (Iran) for technical support for umckalin analysis. The authors express their sincere gratitude to the Research Institute for Biotechnology and Bioengineering, Isfahan University of Technology, Isfahan, Iran.

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Yousefian, Z., Golkar, P. & Mirjalili, M.H. Production Enhancement of Medicinally Active Coumarin and Phenolic Compounds in Hairy Root Cultures of Pelargonium sidoides: The Effect of Elicitation and Sucrose. J Plant Growth Regul 40, 628–641 (2021). https://doi.org/10.1007/s00344-020-10127-y

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