Induction of Callus in Leaf Explants of Crinum americanum L. (Amaryllidaceae)

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Kicia K. P. Gomes- Copeland
Izulmé R. I. Santos
Amanda G. Torres
João V. D. Gomes
Fabrício T. C. de Almeida
Christopher W. Fagg
Sueli M. Gomes
Dâmaris Silveira
Luiz A. Simeoni


Amaryllidaceae include plant species that present alkaloids with analgesic, anti-cancer, anti-bacterial, anti-viral, anti-fungal and anti-malarial activities. Due to this pharmacological value, several species of this family have been widely studied and among them is White lilly, Crinum americanum. The objective of this work was to induce callogenesis on leaf explants of C. americanum cultivated in vitro for future production of alkaloids. Leaf explants were grown on a culture medium (solid) Murashige and Skoog (1962) supplemented with different concentrations and combinations of plant growth regulators, auxin 2,4-dichlorophenoxyacetic acid and cytokinin 6-benzylaminopurine and their effect on callogenesis assessed for percentage oxidation and explants responsive to callus induction. Callus formation started 10 days after hormone inoculation, and within 30 days after inoculation the best callogenesis and callus biomass growth were observed in medium containing 2.5 mg L-1 of 2,4-dichlorophenoxyacetic acid and 10 mg L-1 of 6-benzylaminopurine. The lowest percentage of oxidation was observed on explants cultivated on medium containing 5 mg L-1 of 6-benzylaminopurine and 2.5 mg L-1 of 2,4-dichlorophenoxyacetic acid. The calli obtained were compact and embryogenic. This work contributes not only to future studies on in vitro callogenesis of this species, but also to a possible protocol for the production of alkaloids of interest from cell suspension cultures produced in vitro. This is the first report of callus formation in Crinum americanum explants.

Alkaloids, callogenesis, growth regulators, tissue culture, environmental impact.

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How to Cite
Copeland, K. K. P. G.-, Santos, I. R. I., Torres, A. G., Gomes, J. V. D., Almeida, F. T. C. de, Fagg, C. W., Gomes, S. M., Silveira, D., & Simeoni, L. A. (2020). Induction of Callus in Leaf Explants of Crinum americanum L. (Amaryllidaceae). European Journal of Medicinal Plants, 31(11), 49-56.
Original Research Article


Amaral-Lopes AC, Cavalcanti TB. Brasil Habranthus (Amaryllidaceae) of Brasil. Rodriguesia. 2015;66(1):203-220.

Verdoorn C. The Genus Crinum in Southern Africa. Bothalia. 1973;11(1):27-52.

Fennell CW, van Staden J. Crinum species in traditional and modern medicine. J Ethnopharmacol. 2001;78:15-26.

Nair JJ, van Staden J. Pharmacological and toxicological insights to the South African Amaryllidaceae. Food Chem Toxicol. 2013;62:262-275.

Berkov S, Bastida J, Viladomat F, Codina C. Analysis of galanthamine-type alkaloids by capillary gas chromatography-mass spectrometry in plants. Phytochem Anal. 2008;19(4):285-293.

Liu J, Hu WX, He LF, Ye M, Li Y. Effects of lycorine on HL-60 cells via arresting cell cycle and inducing apoptosis. FEBS Lett. 2004;578(3):245-250.

Bastida J, Lavilla R, Viladomat F. Chemical and biological aspects of Narcissus alkaloids. In: The Alkaloids: Chemistry and Biology. San Diego: Elsevier; 2006.

Takos AM, Rook F. Towards a molecular understanding of the biosynthesis of amaryllidaceae alkaloids in support of their expanding medical use. Int J Mol Sci. 2013; 14(6):11713-11741.

Havelek R, Muthna D, Tomsik P, Kralovec K, Seifrtova M, Cahlikova L, Hostalkova A, Safratova M, Perwein M, Cermakova E, Rezacova M. Anticancer potential of amaryllidaceae alkaloids evaluated by screening with a panel of human cells, real-time cellular analysis and Ehrlich tumor-bearing mice. Chem Biol Interact. 2017;275:121-132.

Ochoa-Villarreal M, Howat S, Hong S, Jang MO, Jin YW, Lee EK, Loake GJ. Plant cell culture strategies for the production of natural products. BMB Rep. 2016;49(3):149-158.

Wilson SA, Roberts SC. Recent advances towards development and commercialization of plant cell culture processes for the synthesis of biomolecules. Plant Biotechnol J. 2012; 10(3):249-268.

Bienaimé C, Melin A, Bensaddek L, Attoumbré J, Nava-Saucedo E, Baltora-Rosset S. Effects of plant growth regulators on cell growth and alkaloids production by cell cultures of Lycopodiella inundata. Plant Cell Tissue Organ Cult. 2015;123(3):523-533.

Hussain A, Ahmed I, Nazir H, Ullah I. Plant Tissue Culture: Current Status and Opportunities. In: Leva A, Rinaldi LMR, editors. Recent Advances in Plant in vitro Culture. Intech Open; 2012.

Amani S, Zarei H, Azar AM, Mashayekh K. Micropropagation of Hippeastrum hybridum. Sci J. 2015;36(4):594-605.

Murashige T, Skoog F. A Revised Medium for Rapid Growth and Bio Assays with Tobacco Tissue Cultures. Physiol Plant. 1962;15(3):473-497.

Ferreira DF. Sisvar: A computer statistical analysis system. Ciênc Agrotec. 2011; 35(6):1039-1042.

Silva TDS, Nepomuceno CF, Borges BPdS, Alvim BFM, De Santana JRF. Multiplicação in vitro de Caesalpinia pyramidalis (Leguminosae). Sitientibus Ser Ciênc Biol. 2013;13:1-6. Portuguese.

Santana-Buzzy N, López-Puc G, Canto-Flick A, Barredo-Pool F, Balam-Uc E, Avile´s-Vinãs S, Solís-Marroquín D, Lecona-Guzmán C, Bello-Bello JJ, Gómez-Uc E, Mijangos-Cortés JO. Ontogenesis of the Somatic Embryogenesis of Habanero Pepper (Capsicum chinense Jacq.). Hort Science. 2009;44(1):113-118.

Menóndez-Yuffá A, García de García E. Morphogenic events during indirect somatic embryogenesis in coffee "Catimor" Protoplasma. 1997(199):208-214.

Thao NTP, Ozaki Y, Okubo H. Callus induction and plantlet regeneration in ornamental Alocasia micholitziana. Plant Cell Tissue Organ Cult. 2003;73(3):285-289.

Sellés M, Viladomat F, Bastida J, Codina C. Callus induction, somatic embryogenesis and organogenesis in Narcissus confusus: Correlation between the state of differentiation and the content of galanthamine and related alkaloids. Plant Cell Rep. 1999;18(7-8):646-651.

Jimenez VM, Bangerth F. Endogenous hormone concentrations and embryogenic callus development in wheat. Plant Cell Tissue Organ Cult. 2001;67:37-46.

Vieira EL, Souza GS, Santos AR, Silva JS. Manual de Fisiologia Vegetal. São Luís: EDUFMA; 2010. Portuguese.

Ptak A, Simlat M, Kwiecień M, Laurain-Mattar D. Leucojum aestivum plants propagated in in vitro bioreactor culture and on solid media containing cytokinins. Eng Life Sci. 2013;13(3):261-270.

Cooke TJ, Poli D, Sztein AE, Cohen JD. Evolutionary patterns in auxin action. Plant Mol Biol. 2002;49(3/4):319-338.

George EF, Hall MA, Klerk GJD. Plant growth regulators II: cytokinins, their analogues and antagonists. In: George EF, Hall MA, Klerk GJD, editors. Plant Propagation by Tissue Culture. Dordrecht: Springer; 2008.

Sahraroo A, Babalar M, Mirjalili MH, Moghaddam MRF, Ebrahimi SN. In-vitro Callus Induction and Rosmarinic Acid Quantification in Callus Culture of Satureja khuzistanica Jamzad (Lamiaceae). Iran J Pharm Res. 2014;13(4):1447-1456.

Ivanov I, Georgiev V, Georgiev M, Ilieva M, Pavlov A. Galanthamine and related alkaloids production by Leucojum aestivum L. shoot culture using a temporary immersion technology. Appl Biochem Biotechnol. 2011;163(2):268-277.

Pilatti FK, Aguiar T, Simões T, Benson EE, Viana AM. In vitro and cryogenic preservation of plant biodiversity in Brazil. In Vitro Cell Dev Biol Plant. 2010;47(1):82-98.