, Volume 70, Issue 2, pp 603–614 | Cite as

A comparative study of biotechnological approaches for producing valuable flavonoids in Prosopis farcta

  • Somaieh Zafari
  • Mohsen SharifiEmail author
  • Najmeh Ahmadian Chashmi
Original Article


The callus and hairy root cultures of Prosopis farcta were established to develop effective strategies to enhance its valuable and medicinally important flavonoid compounds. For callus induction, the hypocotyl, cotyledon and shoot explants were subjected to different plant hormones, naphthalene acetic acid (NAA), benzylaminopurine (BAP), kinetin and dichlorophenoxyacetic acid (2,4-D). Greater callus induction was obtained from hypocotyl explants on MS medium containing 3.0 mg L−1 NAA + 2.0 mg L−1 BAP. With the addition of 0.5 mg L−1 asparagine to this medium, the maximum callus growth was achieved. Hairy root culture of P. farcta was performed using transformation of different explants with strains of Agrobacterium rhizogenes LBA9404, A4, AR15834. The AR15834 strain was more effective for hairy root induction where it caused hairy root formation on 59% of the infected cotyledon explants. We compared profiles of flavonoids isolated from seedling roots, hairy roots, and callus cultures of P. farcta. The colorimetric analysis showed that the content of total flavonoids of hairy roots was 1.54 and 2.52 times higher than in seedling roots and callus, respectively. The presence of flavonoids was verified by LC/MS in positive ion mode. The results showed that flavonoid composition was different in the roots and callus. Naringenin was the major constituent in callus, whereas resveratrol, quercetin and myricetin were the most abundant compounds found in hairy roots. The main objective of this research was to establish hairy roots in P. farcta to synthesize flavonoids at levels comparable to in vitro-grown roots. The present study also opens up a way to further improve the production of pharmaceutically valuable flavonoids and to produce desired metabolites using the hairy root culture system.


Prosopis farcta Hairy root Agrobacterium rhizogenes Callus culture Flavonoids HPLC LC/MS 



Naphthalene acetic acid






Dichlorophenoxyacetic acid


Trichlorophenoxy acetic acid


Dry weight


Cetyltrimethylammonium bromide





This study was financially supported by Tarbiat Modares University, Tehran, Iran.

Author contributions

This research paper was accomplished with the collaboration of authors. SZ performed the experiments, analyzed data and wrote the manuscript. MS designed the study and supervised the study. NAC advised the study.

Supplementary material

10616_2017_143_MOESM1_ESM.docx (706 kb)
Supplementary material 1 (DOCX 705 kb)


  1. Ahmadi-Sakha S, Sharifi M, Niknam V (2016) Bioproduction of phenylethanoid glycosides by plant cell culture of Scrophularia striata Boiss.: from shake-flasks to bioreactor. Plant Cell Tissue Organ Cult 124:275–281. CrossRefGoogle Scholar
  2. Akkol EK, Göger F, Kosar M, Baser HC (2008) Phenolic composition and biological activities of Salvia halophile and Salvia virgata from Turkey. Food Chem 108:942–949. CrossRefGoogle Scholar
  3. Asadollahi K, Abassi N, Afshar N, Alipour M, Asadollahi P (2010) Investigation of the effects of Prosopis farcta plant extract on rat’s aorta. J Med Plants Res 4:142–147. Google Scholar
  4. Batra P, Sharma AK (2013) Anti-cancer potential of flavonoids: recent trends and future perspectives. 3 Biotech 3:439–459. CrossRefGoogle Scholar
  5. Bourgaud F, Gravot A, Milesi S, Gontier E (2001) Production of plant secondary metabolites: a historical perspective. Plant Sci 161:839–851. CrossRefGoogle Scholar
  6. Brahmachari G, Gorai D (2006) Progress in the research on naturally occurring flavones and flavonols: an overview. Curr Org Chem 10:873–898. CrossRefGoogle Scholar
  7. Buendía-González L, Orozco-Villafuerte J, Cruz-Sosa F, Chávez-Ávila VM, Vernon-Carter EJ (2007) Clonal propagation of mesquite tree (Prosopis laevigata Humb. & Bonpl. ex Willd. M.C. Johnston). I. via cotyledonary nodes. In Vitro Cell Dev Biol Plant 43:260–266. CrossRefGoogle Scholar
  8. Caro LA, Polci PA, Lindström LI, Echenique CV, Hernández LF (2002) Micropropagation of Prosopis chilensis (Mol.) Stuntz from young and mature plants. Biocell 26:25–33Google Scholar
  9. Carvalho CHS, Zehr UB, Gunaratna N, Anderson J, Kononowicz HH, Hodges TK, Axtell JD (2004) Agrobacterium-mediated transformation of sorghum: factors that affect transformation efficiency. Genet Mol Biol 27:259–269. CrossRefGoogle Scholar
  10. Chaudhry Z, Afroz A, Rashid H (2007) Effect of variety and plant growth regulators on callus proliferation and regeneration response of three tomato cultivars (Lycopersicon esculentum). Pak J Bot 39:857–869Google Scholar
  11. Estrada-Zúñiga ME, Cruz-Sosa F, Rodrĺguez-Monroy M, Verde-Calvo JR, Vernon-Carter EJ (2009) Phenylpropanoid production in callus and cell suspension cultures of Buddleja cordata Kunth. Plant Cell Tissue Organ Cult 97:39–47. CrossRefGoogle Scholar
  12. Giri A, Narasu ML (2000) Transgenic hairy roots: recent trends and applications. Biotechnol Adv 18:1–22. CrossRefGoogle Scholar
  13. Giri A, Giri CC, Dhingra V, Narasu ML (2001) Enhanced podophyllotoxin production from Agrobacterium rhizogenes transformed cultures of Podophyllum hexandrum. Nat Prod Lett 15:229–235. CrossRefGoogle Scholar
  14. Grzegorczyk-Karolak I, Kuźma L, Wysokińska H (2016) In vitro cultures of Scutellaria alpina as a source of pharmacologically active metabolites. Acta Physiol Plant 38:1–9. CrossRefGoogle Scholar
  15. Gudej J, Tomczyk M (2004) Determination of flavonoids, tannins and ellagic acid in leaves from Rubus L. species. Arch Pharm Res 27:1114–1119. CrossRefGoogle Scholar
  16. Henciya S, Seturaman P, James A, Tsai Y, Nikam R, Wu Y, Dahms H, Chang FR (2017) Biopharmaceutical potentials of Prosopis spp. (Mimosaceae, Leguminosa). J Food Drug Anal 25:187–196. CrossRefGoogle Scholar
  17. Keinänen M, Oldham NJ, Baldwin IT (2001) Rapid HPLC screening of jasmonate induced increases in tobacco alkaloids, phenolics, and diterpene glycosides in Nicotiana attenuata. J Agric Food Chem 49:3553–3558. CrossRefGoogle Scholar
  18. Khan S, Irfan Qureshi M, Kamaluddin Alam T, Abdin MZ (2007) Protocol for isolation of genomic DNA from dry and fresh roots of medicinal plants suitable for RAPD and restriction digestion. Afr J Biotechnol 6:175–178Google Scholar
  19. Khanpour-Ardestani N, Sharifi M, Behmanesh M (2015) Establishment of callus and cell suspension culture of Scrophularia striata Boiss.: an in vitro approach for acteoside production. Cytotechnology 67:475–485. CrossRefGoogle Scholar
  20. Khodashenas M, Keramat B, Emamipoor Y (2015) Callus induction and PLBs production from Levisticum officinale Koch (a wild medicinal plant). J Appl Environ Biol Sci 5:172–180Google Scholar
  21. Khurana S, Venkataraman K, Hollingsworth A, Piche M, Tai TC (2013) Polyphenols: benefits to the cardiovascular system in health and in aging. Nutrients 5:3779–3827. CrossRefGoogle Scholar
  22. Kumar S, Pandey AK (2013) Chemistry and biological activities of flavonoids: an overview. Sci World J 2013:162750. Google Scholar
  23. Lee KH, Morris-Natschke S, Qian K, Dong Y, Yang X, Zhou T, Belding E, Wu SF, Wada K, Akiyama T (2012) Recent progress of research on herbal products used in traditional chinese medicine: the herbs belonging to the divine husbandman’s herbal foundation canon (Shén Nóng Běn Cǎo Jīng). J Tradit Complement Med 2:6–26. CrossRefGoogle Scholar
  24. Masoumian M, Arbakariya A, Syahida A, Maziah M (2011) Effect of precursors on flavonoid production by Hydrocotyle bonariensis callus tissues. Afr J Biotechnol 10:6021–6029. Google Scholar
  25. Mathur S, Shekhawat GS (2013) Establishment and characterization of Stevia rebaudiana (Bertoni) cell suspension culture: an in vitro approach for production of stevioside. Acta Physiol Plant 35:931–939. CrossRefGoogle Scholar
  26. Matkowski A (2008) Plant in vitro culture for the production of antioxidants—a review. Biotechnol Adv 26:548–560. CrossRefGoogle Scholar
  27. Moghadam YA, Piri K, Bahramnejad B, Ghiasvand B (2014) Dopamine production in hairy root cultures of Portulaca oleracea (purslane) using Agrobacterium rhizogenes. J Agric Sci Technol 16:409–420Google Scholar
  28. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Plant Physiol 15:473–497. CrossRefGoogle Scholar
  29. Nandwani D, Ramawat KG (1992) High frequency plantlets regeneration from seedling explants of Prosopis tamarugo. Plant Cell Tissue Organ Cult 29:173–178. CrossRefGoogle Scholar
  30. Park SU, Facchini PJ (2000) Agrobacterium rhizogenes-mediated transformation of opium poppy, Papaver somniferum L., and California poppy, Escholizia californica cham., root cultures. J Exp Bot 51:1005–1016. CrossRefGoogle Scholar
  31. Pawar PK, Maheshwari VL (2004) Agrobacterium rhizogenes mediated hairy root induction in two medicinally important members of family Solanaceae. Indian J Biotechnol 3:414–417Google Scholar
  32. Rai KK, Pandey N, Pandey-Rai S (2014) Effect of 2,4-D on phenolics production and detection of in vitro culture-induced variation through inter-simple sequence repeat and RAPD analysis in Artemisia annua L. Int J Pharm Bio Sci 5:181–193Google Scholar
  33. Rama Krishna D, Shasthree T (2015) Adventitious rooting and proliferation from different explants of Citrullus colocynthis (L.) Schrad an endangered medicinally important cucurbit. Asian J Biotechnol 7:88–95. CrossRefGoogle Scholar
  34. Rao SR, Ravishankar GA (2002) Plant cell cultures: chemical factories of secondary metabolites. Biotechnol Adv 20:101–153. CrossRefGoogle Scholar
  35. Ritchie GA, Short KC, Davey MR (1989) In vitro shoot regeneration from callus, leaf axils and petioles of sugar beet (Beta vulgaris L.). J Exp Bot 40:277–283. CrossRefGoogle Scholar
  36. Robertson S, Narayanan N, Raj Kapoor B (2011) Antitumour activity of Prosopis cineraria (L.) Druce against Ehrlich ascites carcinoma-induced mice. Nat Prod Res 25:857–862. CrossRefGoogle Scholar
  37. Sánchez-Sampedro MA, Fernández-Tárrago J, Corchete P (2009) Elicitation of silymarin in cell cultures of Silybum marianum: effect of subculture and repeated addition of methyl jasmonate. Biotechnol Lett 31:1633–1637. CrossRefGoogle Scholar
  38. Seturaman Prabha D, Dahms HU, Prabha M (2014) Pharmacological potentials of phenolic compounds from Prosopis spp. J Coast Life Med 2:918–924. Google Scholar
  39. Sharifi S, Sattari TN, Zebarjadi A, Majd A, Ghasempour H (2014) The influence of Agrobacterium rhizogenes on induction of hairy roots and ß-carboline alkaloids production in Tribulus terrestris L. Physiol Mol Biol Plants 20:69–80. CrossRefGoogle Scholar
  40. Shekhawat NS, Rathore TS, Singh RP, Deora NS, Rao SR (1993) Factors affecting in vitro clonal propagation of Prosopis cineraria. Plant Growth Regul 12:273–280. CrossRefGoogle Scholar
  41. Shukla S, Gupta S (2010) Apigenin: a promising molecule for cancer prevention. Pharm Res 27:962–978. CrossRefGoogle Scholar
  42. Smith MAL, Kobayashi H, Gawienowski M, Briskin DP (2002) An in vitro approach investigate medicinal chemical synthesis by three herbal plants. Plant Cell Tissue Organ Cult 70:105–111. CrossRefGoogle Scholar
  43. Staszków A, Swarcewicz B, Banasiak J, Muth D, Jasiński M, Stobiecki M (2011) LC/MS profiling of flavonoid glycoconjugates isolated from hairy roots, suspension root cell cultures and seedling roots of Medicago truncatula. Metabolomics 7:604–613. CrossRefGoogle Scholar
  44. Taghipour F, Janalizadeh N, Eshrati M, Hassanzadeh T, Huyop F (2013) Callus induction and shoot organogenesis in two sugar beet (Bera vulgaris L.). breeding lines in vitro cultured. Biotechnology 12:168–178. CrossRefGoogle Scholar
  45. Toker G, Memisoglu M, Toker MC, Yesilada E (2003) Callus formation and cucurbitacin B accumulation in Echallium elaterium callus cultures. Fitoterapia 74:618–623. CrossRefGoogle Scholar
  46. Trejo-Espino JL, Rodríguez-Monroy M, Vernon-Carter EJ, Cruz-Sosa F (2011) Establishment and characterization of Prosopis laevigata (Humb. & Bonpl. ex Willd) M.C. Johnst. cell suspension culture: a biotechnology approach for mesquite gum production. Acta Physiol Plant 33:1687–1695. CrossRefGoogle Scholar
  47. Wang B, Zhang G, Zhu L, Chen L, Zhang Y (2006) Genetic transformation of Echinacea purpurea with Agrobacterium rhizogenes and bioactive ingredient analysis in transformed cultures. Colloids Surf B Biointerfaces 53:101–104. CrossRefGoogle Scholar
  48. Zafari S, Sharifi M, Ahmadian-Chashmi N, Mur LAJ (2016) Modulation of Pb-induced stress in Prosopis shoots through an interconnected network of signaling molecules, phenolic compounds and amino acids. Plant Physiol Biochem 99:11–20. CrossRefGoogle Scholar
  49. Zafari S, Sharifi M, Ahmadian-Chashmi N (2017a) Nitric oxide production shifts metabolic pathways toward lignification to alleviate Pb stress in Prosopis farcta. Environ Exp Bot 141:41–49. CrossRefGoogle Scholar
  50. Zafari S, Sharifi M, Mur LAJ, Ahmadian-Chashmi N (2017b) Favouring NO over H2O2 production will increase Pb tolerance in Prosopis farcta via altered primary metabolism. Ecotoxicol Environ Saf 142:293–302. CrossRefGoogle Scholar
  51. Zawawi DD, Jaafar H, Ali AM (2013) Effects of 2,4-D and kinetin on callus induction of Barringtonia racemosa leaf and endosperm explants in different types of basal media. Asian J Plant Sci 12:21–27. CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2018

Authors and Affiliations

  • Somaieh Zafari
    • 1
  • Mohsen Sharifi
    • 1
    Email author
  • Najmeh Ahmadian Chashmi
    • 2
  1. 1.Department of Plant Biology, Faculty of Biological SciencesTarbiat Modares UniversityTehranIran
  2. 2.Department of Biology, Faculty of Basic SciencesUniversity of MazandaranBabolsarIran

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