Advertisement

Ocimum basilicum suspension culture as resource for bioactive triterpenoids: yield enrichment by elicitation and bioreactor cultivation

  • Pallavi Pandey
  • Sailendra Singh
  • Suchitra BanerjeeEmail author
Original Article
  • 100 Downloads

Abstract

The competence of Ocimum basilicum suspension culture for the production of pharmacologically distinguished triterpenoids had been explored to meet their escalating demand with inequitable natural resources. The production of three pentacyclic triterpenes—betulinic acid (BA), ursolic acid (UA) and oleanolic acid (OA), could be achieved for the first time in the O. basilicum suspension culture, together with the prevalently occurring rosmarinic acid (RA). The production of RA was most abundant (15.73 ± 0.28 mg/g DW) followed by that of BA (14.63 ± 0.21 mg/g DW), UA (4.71 ± 0.23 mg/g DW) and OA (0.91 ± 0.02 mg/g DW) in reducing order on their respective optimum cultivation period. Compared to the in-vivo control leaves, the suspension cell revealed almost 3.25-, 2.89-, and 1.79-folds higher OA, UA and RA contents, respectively. The BA synthesizing power of suspension cells exemplified notable trend as its presence in the control leaves remained obscured. By and large, cultivation in a 10 l bioreactor and elicitation with methyl jasmonate in bioreactor and shake flask improved the cumulative productivities of the targeted metabolites with reciprocated divergence in the amounts of the two isomers, i.e., UA and OA. The overall findings of the present study bear special merit in realising the global demand of these multiple terpenoids in a single culture system of O. basilicum, which remained unrevealed so far.

Key Message

First report towards the in vitro production of multiple pharmacologically distinguished, high demand triterpenoids—betulinic acid (BA), ursolic acid (UA) and oleanolic acid (OA), along with rosmarinic acid (RA) in suspension culture of Ocimum basilicum. Notable yield enhancement through bioreactor cultivation and strategic elicitation with methyl jasmonate in shake flask and bioreactor added additional merit to this study in realizing the global demand of these multiple terpenoids in a single culture system of O. basilicum.

Keywords

Bioreactor cultivation Cell suspension Elicitation Labiateae Triterpenoids 

Notes

Acknowledgements

The authors wish to express their sincere thanks to the Director, CSIR-CIMAP, for providing the facilities to carry out this research. Financial supports from Department of Science and Technology (DST, New Delhi, India) and the National Academy of Sciences, India (NASI- Allahabad) are gratefully acknowledged. Sincere gratitude is also due to the Editor and Reviewers of this manuscript for their critical comments and knowledgeable suggestion that upgraded this work through incorporation of the “bioreactor-elicitation” combined study. This study was initially funded by CSIR network project (BSC-0203).

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest.

References

  1. Bhuvaneshwari K, Gokulanathan A, Jayanthi M, Govindasamy V, Milella L, Lee S, Yang DC, Girija S (2016) Can Ocimum basilicum L. and Ocimum tenuiflorum L. in vitro culture be a potential source of secondary metabolites? Food Chem 194:55–60CrossRefGoogle Scholar
  2. Bolta Z, Baricevic D, Bohanec B, Andrensek S (2000) Preliminary investigation of ursolic acid in cell suspension culture of Salvia officinalis L. Plant Cell Tissue Organ Cult 62:63–75CrossRefGoogle Scholar
  3. Bora KS, Arora S, Shri R (2011) Role of Ocimum basilicum L. in prevention of ischemia and reperfusion-induced cerebral damage, and motor dysfunctions in mice brain. J Ethnopharmacol 137:1360–1365CrossRefGoogle Scholar
  4. Dzubak P, Hajduch M, Vydra D, Hustova A, Kvasnica M, Biedermann D, Markova L, Urban M, Sarek J (2006) Pharmacological activities of natural triterpenoids and their therapeutic implications. Nat Prod Rep 23:394–411CrossRefGoogle Scholar
  5. Feria-Romero I, Lazo E, Ponce-Noyola T, Cerda-Garcia-Rojas CM, Ramos-Valdivia AC (2005) Induced accumulation of oleanolic acid and ursolic acid in cell suspension cultures of Uncaria tomentosa. Biotechnol Lett 27:839–843CrossRefGoogle Scholar
  6. Georgiev M, Pavlov A, Ilieva M (2004) Rosmarinic acid production by Lavandula vera MM cell suspension: the effect of temperature. Biotechnol Lett 26:855–856CrossRefGoogle Scholar
  7. Georgiev MI, Eibl R, Zhong JJ (2013) Hosting the plant cells in vitro: recent trends in bioreactors. Appl Microbiol Biotechnol 97:3787–3800CrossRefGoogle Scholar
  8. Ghaffari-Moghaddam M, Ahmad FBH, Samadzadeh-Kermani A (2012) Biological activity of betulinic acid: a review. Pharmacol Pharm 3:119–123CrossRefGoogle Scholar
  9. Gupta R, Pandey P, Singh S, Singh DK, Saxena A, Luqman S, Bawankule DU, Banerjee S (2016) Advances in Boerhaavia diffusa hairy root technology: a valuable pursuit for identifying strain sensitivity and up-scaling factors to refine metabolite yield and bioactivity potentials. Protoplasma 253:1145–1158CrossRefGoogle Scholar
  10. Haas C, Hengelhaupt KC, Kummritz S, Bley T, Pavlov A, Steingroewer J (2014) Salvia suspension cultures as production systems for oleanolic and ursolic acid. Acta Physiol Plant 36:2137CrossRefGoogle Scholar
  11. Hajati RJ, Payamnoor V, Chashmi NA, Bezdi KG (2018) Improved accumulation of betulin and betulinic acid in cell suspension culture of Betula pendula Roth by abiotic and biotic elicitors. Prep Biochem Biotechnol.  https://doi.org/10.1080/10826068.2018.1514514 Google Scholar
  12. Hakkim FL, Kalyani S, Essa M, Girija S, Song H (2011) Production of rosmarinic acid in Ocimum sanctum (L.) cell suspension cultures by the influence of growth regulators. Int J Biol Med Res 2:1158–1161Google Scholar
  13. Isah T, Umar S, Mujib A, Sharma MP, Rajasekharan PE, Zafar N, Frukh A (2018) Secondary metabolism of pharmaceuticals in the plant in vitro cultures: strategies, approaches, and limitations to achieving higher yield. Plant Cell Tissue Organ Cult 132:239–265CrossRefGoogle Scholar
  14. Jager S, Trojan H, Kopp T, Laszczyk MN, Scheffler A (2009) Pentacyclic triterpene distribution in various plants rich sources for a new group of multi-potent plant extracts. Molecules 14:2016–2031CrossRefGoogle Scholar
  15. Jamil SZMR, Rohani ER, Baharum SN, Noor NM (2018) Metabolite profiles of callus and cell suspension cultures of mangosteen. 3 Biotech 8:322CrossRefGoogle Scholar
  16. Karam NS, Jawad FM, Arikat NA, Shibli RA (2003) Growth and rosmarinic acid accumulation in callus, cell suspension, and root cultures of wild Salvia fruticosa. Plant Cell Tissue Organ Cult 73:117–121CrossRefGoogle Scholar
  17. Khojasteh A, Mirjalili MH, Palazon J, Eibl R, Cusido RM (2016) Methyl jasmonate enhanced production of rosmarinic acid in cell cultures of Satureja khuzistanica in a bioreactor. Eng Life Sci 16:740–749CrossRefGoogle Scholar
  18. Kim GD, Park YS, Jin YH, Park CS (2015) Production and applications of rosmarinic acid and structurally related compounds. Appl Microbiol Biotechnol 99:2083–2092CrossRefGoogle Scholar
  19. Kintzios S, Makri O, Panagiotopoulos E, Scapeti M (2003) In vitro rosmarinic acid accumulation in sweet basil (Ocimum basilicum L). Biotechnol Lett 25:405–408CrossRefGoogle Scholar
  20. Kintzios S, Kollias H, Straitouris E, Makri O (2004) Scale-up micropropagation of sweet basil (Ocimum basilicum L.) in an airlift bioreactor and accumulation of rosmarinic acid. Biotechnol Lett 26:521–523CrossRefGoogle Scholar
  21. Krzyzanowska J, Czubacka A, Pecio L, Przybys M, Doroszewska T, Stochmal A, Oleszek W (2012) The effects of jasmonic acid and methyl jasmonate on rosmarinic acid production in Mentha × piperita cell suspension cultures. Plant Cell Tissue Organ Cult 108:73–81CrossRefGoogle Scholar
  22. Li H, Fu Y, Sun H, Zhang Y, Lan X (2017) Transcriptomic analyses reveal biosynthetic genes related to rosmarinic acid in Dracocephalum tanguticum. Sci Rep 7:74CrossRefGoogle Scholar
  23. Liu J (2005) Oleanolic acid and ursolic acid: research perspectives. J Ethnopharmacol 100:92–94CrossRefGoogle Scholar
  24. Luo H, Zhu Y, Song J, Xu L, Sun C, Zhang X, Xu Y, He L, Sun W, Xu H, Wang B (2014) Transcriptional data mining of Salvia miltiorrhiza in response to methyl jasmonate to examine the mechanism of bioactive compound biosynthesis and regulation. Physiol Plant 152:241–255CrossRefGoogle Scholar
  25. Makri O, Kintzios S (2007) Ocimum sp. (Basil): botany, cultivation, pharmaceutical properties, and biotechnology. J Herbs Spices Med Plants 13:123–150CrossRefGoogle Scholar
  26. Martínez VVM, Estrada-Soto SE, Arellano-García JD, Rivera-Leyva JC, Castillo-Espana P, Flores AF, Cardoso-Taketa AT, Perea-Arango I (2017) Methyl jasmonate and salicylic acid enhanced the production of ursolic and oleanolic acid in callus cultures of Lepechinia Caulescens. Phcog Mag 13:S886–S889Google Scholar
  27. Marzouk AM (2009) Hepatoprotective triterpenes from hairy root cultures of Ocimum basilicum L. Z Naturforsch 64c:201–209CrossRefGoogle Scholar
  28. Mathew R, Sankar PD (2012) Effect of methyl jasmonate and chitosan on growth characteristics of Ocimum basilicum L., Ocimum sanctum L. and Ocimum gratissimum L. cell suspension cultures. Afr J Biotechnol 11:4759–4766Google Scholar
  29. Misra RC, Maiti P, Chanotiya CS, Shanker K, Ghosh S (2014) Methyl jasmonate-elicited transcriptional responses and pentacyclic triterpene biosynthesis in sweet basil. Plant Physiol 164:1028–1044CrossRefGoogle Scholar
  30. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol Plant 15:473–497CrossRefGoogle Scholar
  31. 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 16:1227–1252CrossRefGoogle Scholar
  32. Norrizah JS, Suhaimi Y, Rohaya A, Roslan NAR (2012) Ursolic acid and oleanolic acid productions in elicited cell suspension cultures of Hedyotis corymbosa. Biotechnology 11:238–242CrossRefGoogle Scholar
  33. Pandey H, Pandey P, Singh S, Gupta R, Banerjee S (2015) Production of anti-cancer triterpene (betulinic acid) from callus cultures of different Ocimum species and its elicitation. Protoplasma 252:647–655CrossRefGoogle Scholar
  34. Pavlov A, Georgiev M, Panchev I, Ileva M (2005a) Optimization of rosmarinic acid production by Lavandula vera MM plant cell suspension in a laboratory bioreactor. Biotechnol Prog 21:394–396CrossRefGoogle Scholar
  35. Pavlov AI, Georgiev MI, Ilieva MP (2005b) Production of rosmarinic acid by Lavandula vera MM cell suspension in bioreactor: effect of dissolved oxygen concentration and agitation. World J Microbiol Biotechnol 21:389–392CrossRefGoogle Scholar
  36. Pereira PS, Ticli FK, França SDC, Breves CMDS, Lourenço MV (2007) Enhanced triterpene production in Tabernaemontana catharinensis cell suspension cultures in response to biotic elicitors. Quim Nova 30:1849–1852CrossRefGoogle Scholar
  37. Petersen M, Hausler E, Meinhard J, Karwatzki B, Gartlowski C (1994) The biosynthesis of rosmarinic acid in suspension cultures of Coleus blumei. Plant Cell Tissue Organ Cult 38:171–179CrossRefGoogle Scholar
  38. Qamar KA, Dar A, Siddiqui BS (2010) Anticancer activity of Ocimum basilicum and the effect of ursolic acid on the cytoskeleton of MCF-7 human breast cancer cells. Lett Drug Design Dis 7:726–736CrossRefGoogle Scholar
  39. Rahman RAIA, El-Wakil HED, Abdelsalam NR, Elsaadany RMA (2015) In-vitro production of rosmarinic acid from basil (Ocimum basilicum L.) and Lemon Balm (Melissa officinalis L.). Middle East J Appl Sci 5:47–51Google Scholar
  40. Santos-Gomes PC, Seabra RM, Andrade PB, Fernandes-Ferreira M (2003) Determination of phenolic antioxidant compounds produced by calli and cell suspensions of sage (Salvia officinalis L.). J Plant Physiol 160:1025–1032CrossRefGoogle Scholar
  41. Singh A, Dwivedi P (2018) Methyl jasmonate and salicylic acid as potent elicitors for secondary metabolite production in medicinal plants: a Review. J Pharma Phytochem 7:750–757Google Scholar
  42. Sivanandhan G, Selvaraj N, Ganapathi A, Manickavasagam M (2014) Enhanced biosynthesis of withanolides by elicitation and precursor feeding in cell suspension culture of Withania somnifera (L.) Dunal in shake flask culture and bioreactor. PLoS ONE 9(8):e104005CrossRefGoogle Scholar
  43. Song H, Kumar P, Arivazhagan G, Sang-I L, Yoon HM, Kim IH, Kwon HJ, Kim JM, Hakkim L (2012) Antioxidant property of leaves and calluses extracts of in-vitro grown 5 different Ocimum species. J Plant Biotechnol 39:146–153CrossRefGoogle Scholar
  44. Srivastava P, Kasoju N, Bora U, Chaturvedi R (2010) Accumulation of betulinic, oleanolic and ursolic acids in in vitro cell cultures of Lantana camara L. and their significant cytotoxic effects on HeLa cell lines. Biotechnol Bioproc Eng 15:1038–1046CrossRefGoogle Scholar
  45. Swamy MK, Sinniah UR, Ghasemzadeh A (2018) Anticancer potential of rosmarinic acid and its improved production through biotechnological interventions and functional genomics. Appl Microbiol Biotechnol 102:7775–7793CrossRefGoogle Scholar
  46. Szabo E, Thelen A, Petersen M (1999) Fungal elicitor preparations and methyl jasmonate enhance rosmarinic acid accumulation in suspension cultures of Coleus blumei. Plant Cell Rep 18:485–489CrossRefGoogle Scholar
  47. Tada H, Murakami Y, Omoto T, Shimomuro K, Ishimaru K (1996) Rosmarinic acid and related phenolics in hairy root cultures of Ocimum basilicum. Phytochemistry 42:431–434CrossRefGoogle Scholar
  48. Ulbrich B, Weisner W, Arens H (1985) Large scale production of rosmarinic acid from plant cell cultures of Coleus blumei Benth. In: Neumann KH, Barz W, Reinhard E (eds) Primary and secondary metabolism of plant cell cultures. Springer, Berlin, pp 293–303CrossRefGoogle Scholar
  49. Wang JW, Xia ZH, Chu JH, Tan RX (2004) Simultaneous production of anthocyanin and triterpenoids in suspension cultures of Perilla frutescens. Enzyme Microb Technol 34:651–656CrossRefGoogle Scholar
  50. Wang W, Zhang ZY, Zhong JJ (2005) Enhancement of ginsenoside biosynthesis in high density cultivation of Panax notoginseng cells by various strategies of methyl jasmonate elicitation. Appl Microbiol Biotechnol 67:752–758CrossRefGoogle Scholar
  51. Wang W, Zhao Z, Xu Y, Qian X, Zhong JJ (2006) Efficient induction of ginsenoside biosynthesis and alteration of ginsenoside heterogeneity in cell cultures of Panax notoginseng by using chemically synthesized 2-hydroxyethyl jasmonate. Appl Microbiol Biotechnol 70:298–307CrossRefGoogle Scholar
  52. Wang QJ, Lei XY, Zheng LP, Wang JW (2017) Molecular characterization of an elicitor-responsive 3-hydroxy-3-methylglutaryl coenzyme A reductase gene involved in oleanolic acid production in cell cultures of Achyranthes bidentata. Plant Growth Regul 81:335–343CrossRefGoogle Scholar
  53. Wiktorowska E, Dlugosz M, Janiszowska W (2010) Significant enhancement of oleanolic acid accumulation by biotic elicitors in cell suspension cultures of Calendula officinalis L. Enzyme Microb Technol 46:14–20CrossRefGoogle Scholar
  54. Xie Z, Kapteyn J, Gang DR (2008) A systems biology investigation of the MEP/terpenoid and shikimate/phenylpropanoid pathways points to multiple levels of metabolic control in sweet basil glandular trichomes. Plant J 54:349–361CrossRefGoogle Scholar
  55. Yin Z, Shangguan X, Chen J, Zhao Q, Li D (2013) Growth and triterpenic acid accumulation of Cyclocarya paliurus cell suspension cultures. Biotechnol Bioproc E 18:606–614CrossRefGoogle Scholar
  56. Yue W, Ming QL, Lin B, Rahman K, Zheng CJ, Han T, Qin LP (2015) Medicinal plant cell suspension cultures: pharmaceutical applications and high-yielding strategies for the desired secondary metabolites. Crit Rev Biotechnol 20:1–18Google Scholar
  57. Zahra K, Khan MA, Iqbal F (2015) Oral supplementation of Ocimum basilicum has the potential to improve the locomotory, exploratory, anxiolytic behavior and learning in adult male albino mice. Neurol Sci 36:73–78CrossRefGoogle Scholar
  58. Zhang ZY, Zhong JJ (2004) Scale-up of centrifugal impeller bioreactor for hyperproduction of ginseng saponin and polysaccharide by high-density cultivation of Panax notoginseng cells. Biotechnol Prog 20:1076–1081CrossRefGoogle Scholar
  59. Zhong JJ, Zhang ZY (2005) High density cultivation of Panax notoginseng cell cultures with methyl jasmonate elicitation in a centrifugal-impeller bioreactor. Eng Life Sci 5:471–474CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Pallavi Pandey
    • 1
  • Sailendra Singh
    • 1
  • Suchitra Banerjee
    • 1
    Email author
  1. 1.Department of Plant BiotechnologyCentral Institute of Medicinal and Aromatic Plants (CSIR-CIMAP)LucknowIndia

Personalised recommendations