European Journal of Plant Pathology

, Volume 153, Issue 1, pp 37–49 | Cite as

Study of sodium 3-hydroxycoumarin as inhibitors in vitro, in vivo and in silico of Moniliophthora perniciosa fungus

  • Priscila de Andrade Gonçalves
  • Manoelito Coelho dos Santos Junior
  • Catiane do Sacramento Sousa
  • Aristóteles Góes-Neto
  • Edna Dora Martins Newman Luz
  • Virgínia Oliveira Damaceno
  • Ana Rosa Rocha Niella
  • José Maria Barbosa Filho
  • Sandra Aparecida de AssisEmail author


The inhibition of the fungus Moniliophthora perniciosa, which is the causal agent of witches’ broom disease in Theobroma cacao L., is the main focus of the present study. The use of phytoalexins has provided disease control that is as efficient as control with the use of fungicides, with the advantage of not harming humans and the environment. In this sense, the objective was to evaluate the action of 3-hydroxycoumarin compared with the standard defence activator acibenzolar-S-methyl (Bion® 500 WG) and systemic fungicide tebuconazole (Folicur® 200 CE), and study the possible mechanisms of action. Initially, basidiospore germination inhibition assays (in vitro) were performed using four different concentrations of 3-hydroxycoumarin. The result was 100% inhibition at the concentration of 1000 ppm. Thereafter, this substance was used in four different treatments (in vivo) of cacao seedlings of the SIC-23 genotype, with regard to the order of application. The data analysis showed greater inhibition when 3-hydroxycoumarin was applied after inoculation (TAI test), and suggested a curative effect on Theobroma cacao seedlings. In silico, it was used the enzyme chitin synthase because this action as key enzyme in chitin biosynthetic pathway. The in silico tests show that 3-hydroxycoumarin inhibited the production of chitin synthase by the fungus. Therefore, the present study identifies a potential inhibitor of M. perniciosa and suggests the best application methodology.


Witches’ broom Theobroma cacao Chemical control Basidiospores germination TAI test Curative effect 


Compliance with ethical standards

Conflict of interest

The authors state that there are no conflicts of interest.


  1. Aime, M. C., & Phillips-Mora, W. (2005). The causal agents of witches’ broom and frosty pod rot of cacao (chocolate, Theobroma cacao) form a new lineage of Marasmiaceae. Mycologia, 97, 1012–1022.Google Scholar
  2. Al-Amiery, A. A., Al-Bayati, R. I. H., Saour, K. Y., & Radi, M. F. (2011). Cytotoxicity, antioxidant, and antimicrobial activities of novel 2-quinolone derivatives derived from coumarin. Research on Chemical Intermediates, 38, 559–569.CrossRefGoogle Scholar
  3. Al-Amiery, A. A., Kadhum, A. A. H., & Mohamad, A. B. (2012). Antifungal activities of new coumarins. Molecules, 17, 5713–5723.CrossRefGoogle Scholar
  4. Ampasala, D. R., Zheng, S., Zhang, D., Ladd, T., Doucet, D., Krell, P. J., Retnakaran, A., & Feng, Q. (2011). An epidermis-specific chitin synthase cdna in Choristoneura fumiferana: Cloning, characterization, developmental and hormonal-regulated expression. Archives of Insect Biochemistry and Physiology, 76(2), 83–96.CrossRefGoogle Scholar
  5. Atmaka, M., Bilgin, H. M., Obay, B. D., Diken, H., Kelle, M., & Kale, E. (2011). The hepatoprotective effect of coumarin and coumarin derivatives on carbon tetrachloride-induced hepatic injury by anti-oxidative activities in rats. Journal of Physiology and Biochemistry, 67, 569–576.CrossRefGoogle Scholar
  6. Bansal, Y., Sethi, P., & Bansal, G. (2013). Coumarin: A potential nucleus for anti-inflammatory molecules. Medicinal Chemistry Research, 22, 3049–3060.CrossRefGoogle Scholar
  7. Behr, J. B. (2003). Chitin synthase as an antifungal target: Recent advances. Current Medicinal Chemistry, 2, 173–189.Google Scholar
  8. Carpinella, M. C., Ferrayoli, C. G., & Palacios, S. M. (2005). Antifungal synergistic effect of scopoletin, a hydroxycoumarin isolated from Melia azedarach L. fruits. Agricultural and Food Chemistry, 53, 2922–2927.CrossRefGoogle Scholar
  9. Case, D. A., Darden, T. A., Cheatham III, T. E., Simmerling, C. L., Wang, J., Duke, R. E., Luo, R., Walker, R. C., Zhang, W., Merz, K. M., Roberts, B., Hayik, S., Roitberg, A., Seabra, G., Swails, J., Goets, A. W., Kolossváry, I., Wong, K. F., Paesani, F., Vanicek, J., Wolf, R. M., Liu, J., Wu, X., Brozell, S. R., Steinbrecher, T., Gohlke, H., Cai, Q., Ye, X., Wang, J., Hsieh, M. J., Cui, G., Roe, D. R., Mathews, D. H., Seetin, M. G., Salomon-Ferrer, R., Sagui, C., Babin, V., Luchko, T., Gusarov, S., Kovalenko, A., Kollman, P. A. (2012), AMBER 12, University of California, San Francisco.Google Scholar
  10. Dhanavade, M. J., Jalkute, C. B., Ghosh, J. S., & Sonawane, K. D. (2011). Study antimicrobial activity of lemon (Citrus Lemon L.) peel extract. British Journal of Pharmacology and Toxicology, 2, 119–122.Google Scholar
  11. Duo-Chuan, L. (2006). Review of fungal chitinases. Mycopathologia, 161, 345–360.CrossRefGoogle Scholar
  12. Ferreira, S. Z., Carneiro, H. C., Lara, H. A., Alves, R. B., Resende, J. M., Oliveira, H. M., & Freitas, R. P. (2015). Synthesis of a new peptide–coumarin conjugate: A potential agent against Cryptococcosis. ACS Medicinal Chemistry Letters, 6, 271–275.CrossRefGoogle Scholar
  13. Frias, G. A., Purdy, L. H., & Schmidt, R. A. (1995). An inoculation method for evaluate resistance of cocoa to Crinipellis perniciosa. Plant Disease, 79, 787–791.CrossRefGoogle Scholar
  14. Frisch, M. J., Trucks, G. W., Schlegel, H. B., Scuseria, G. E., Robb, M. A., Cheeseman, J. R., Scalmani, G., Barone, V., Petersson, G. A., Nakatsuji, H., Li, X., Caricato, M., Marenich, A., Bloino, J., Janesko, B. G., Gomperts, R., Mennucci, B., Hratchian, H. P., Ortiz, J. V., Izmaylov, A. F., Sonnenberg, J. L., Williams-Young, D., Ding, F., Lipparini, F., Egidi, F., Goings, J., Peng, B., Petrone, A., Henderson, T., Ranasinghe, D., Zakrzewski, V. G., Gao, J., Rega, N., Zheng, G., Liang, W., Hada, M., Ehara, M., Toyota, K., Fukuda, R., Hasegawa, J., Ishida, M., Nakajima, T., Honda, Y., Kitao, O., Nakai, H., Vreven, T., Throssell, K., Montgomery, Jr., J. A., Peralta, J. E., Ogliaro, F., Bearpark, M., Heyd, J. J., Brothers, E., Kudin, K. N., Staroverov, V. N., Keith, T., Kobayashi, R., Normand, J., Raghavachari, K., Rendell, A., Burant, J. C., Iyengar, S. S., Tomasi, J., Cossi, M., Millam, J. M., Klene, M., Adamo, C., Cammi, R., Ochterski, J. W., Martin, R. L., Morokuma, K., Farkas, O., Foresman, J. B., and Fox, D. J., Gaussian, Inc., Wallingford CT, 2016.Google Scholar
  15. Godoy, M. F. P., Victor, S. R., Bellini, A. M., Guerreiro, G., Rocha, W. C., Bueno, O. C., Hebling, M. J. A., Bacci Jr., M., Silva, M. F. G. F., Vieira, P. C., Fernandes, J. B., & Pagnocca, F. C. (2005). Inhibition of the symbiotic fungus of leaf-cutting ants by coumarins. Journal of the Brazilian Chemical Society, 16, 669–672.CrossRefGoogle Scholar
  16. Gramacho, K. P., Luz, E. D. M. N., Silva, F. S., Lopes, U. V., Pires, J. L., & Pereira, L. (2016). Pathogenic variability of Moniliophthora perniciosa in three agroecological zones of the cacao region of Bahia, Brazil. Crop Breeding and Applied Biotechnology, 16, 7–13.CrossRefGoogle Scholar
  17. Johann, S., Mendes, B. G., Missau, F. C., Resende, M. A., & Pizzolatti, M. G. (2011). Antifungal activity of five species of Polygala. Brazilian Journal of Microbiology, 42, 1065–1075.CrossRefGoogle Scholar
  18. Karatas, M. O., Olgundeniz, B., Günal, S., Özdemir, I., Alici, B., & Çetinkaya, E. (2016). Synthesis, characterisation and antimicrobial activities of novel silver(I) complexes with coumarin substituted N-heterocyclic carbene ligands. Bioorganic & Medicinal Chemistry, 24, 643–650.CrossRefGoogle Scholar
  19. Kasumbwe, K., Venugopala, K. N., Mohanlall, V., & Odhav, B. (2014). Antimicrobial and antioxidant activities of substituted halogenated coumarins. Journal of Medicinal Plant Research, 5, 274–281.CrossRefGoogle Scholar
  20. Kerrigan, J. E. (2009). Amber 9.0 Drug/DNA complex. Piscataway, New Jersey, p. 17.Google Scholar
  21. Krumrine, J., Raubacher, F., Brooijmans, N., & Kuntz, I. (2003). Principles and methods of docking and ligand design. In P. E. Bourne & W. Helge (Eds.), Structural Bioinformatics (pp. 443–476). New Jersey: John Wiley & Sons.Google Scholar
  22. Lagorce, A., Le Berre-Anton, V., Aguilar-Uscanga, B., Martin-Yken, H., Dagkessamanskaia, A., & François, J. (2002). Activation of the chitin synthesis pathway in response to cell-wall defects in Saccharomyces cerevisiae. European Journal of Biochemistry, 269, 1697–1707.CrossRefGoogle Scholar
  23. Leal, L. K. A. M., Ferreira, A. A. G., Bezerra, G. A., Matos, F. J. A., & Viana, G. S. B. (2000). Anti-nociceptive, anti-inflammatory and bronchodilator activities of Brazilian medicinal plants containing coumarin: A comparative study. Journal of Ethnopharmacology, 70, 151–150.CrossRefGoogle Scholar
  24. Liu, X., Li, F., Li, D., Ma, E., Zhang, W., Zhu, K. Y., & Zhang, J. (2013). Molecular and functional analysis of UDP-N-Acetylglucosamine pyrophosphorylases from the migratory locust, locusta migratoria. PLoS One, 8(8), e71970. Published online 2013 Aug 19. Scholar
  25. Meinhardt, L. W., Rincones, J., Bailey, B. A., Aime, M. C., Griffith, G. W., & Zhang, D. (2008). Moniliophthora perniciosa, the causal agent of witches’ broom disease of cacao: what’s new from this old foe? Molecular Plant Pathology, 9, 577–588.CrossRefGoogle Scholar
  26. Miyano, D. M., Lima, T., Simões, F. R., La-Scalea, M. A., Oliveira, h. P. M., & Codognoto, L. (2014). Electrochemical study of simple coumarin and its determination in aqueous infusion of Mikania glomerata. Journal of Brazilian Chemical Society, 25, 602–609.Google Scholar
  27. Mouri, T., Yano, T., Kochi, S., Ando, T., & Hori, M. (2005). Synthesis and antifungal activity of new 3,4,7-tri-substituted coumarins. Journal of Pesticide Science, 30, 209–213.CrossRefGoogle Scholar
  28. Rehman, S. U., Chohan, Z. H., Gulnaz, F., & Supuran, C. T. (2005). In-vitro antibacterial, antifungal and cytotoxic activities of some coumarins and their metal complexes. Journal of Enzyme Inhibition and Medicinal Chemistry, 20, 333–340.CrossRefGoogle Scholar
  29. Rino, J. P., & Studart, N. (2001). Um potencial de interação para o estudo de materiais e simulações por dinâmica molecular. Química Nova, 24, 838–845.CrossRefGoogle Scholar
  30. Roncero, C. (2002). The genetic complexity of chitin synthesis in fungi. Current Genetics, 41, 367–378.CrossRefGoogle Scholar
  31. Scarpari, l. M., Meinhardt, l. W., Mazzafera, P., Pomella, A. W. V., Schiavinato, M. A., Cascardo, J. C. M., & Pereira, G. A. G. (2005). Biochemical changes during the development of witches’ broom: The most important disease of cocoa in Brazil caused by Crinipellis perniciosa. Journal of Experimental Botany, 56, 856–877.CrossRefGoogle Scholar
  32. Shi, Y., Zhou, C. H., Zhou, X. D., Geng, R. X., & Ji, Q. G. (2011). Synthesis and antimicrobial evaluation of coumarin-based benzotriazoles and their synergistic effects with chloromycin and fluconazole. Acta Pharmaceutica Sinica, 46, 798–810.Google Scholar
  33. Singh, L. K., Priyanka, Singh, V., & Katiyar, D. (2015). Design, synthesis and biological evaluation of some new coumarin derivatives as potential antimicrobial agents. Medicinal Chemistry, 11, 128–134.CrossRefGoogle Scholar
  34. Souza, C. S., Oliveira, B. M., Costa, G. G. L., Schriefer, A., Schnadelbach, A. S., Uetanabaro, A. P. T., Pirovani, C. P., Pereira, G. A., Taranto, A. G., Cascardo, J. C. M., & Goes-Neto, A. (2009). Identification and characterization of a class III chitin synthase gene of Monilliophthora perniciosa, the fungus that causes witches' broom disease of cacao. Journal of Microbiology, 47, 431–440.CrossRefGoogle Scholar
  35. Stein, A. C., Alvarez, S., Avancini, C., Zacchino, S., & von Poser, G. (2006). Antifungal activity of some coumarins obtained from species of Pterocaulon (Asteraceae). Journal of Ethnopharmacology, 107, 95–98.CrossRefGoogle Scholar
  36. Stewart, J. J. P. (2007). Optimization of parameters for semi-empirical methods V: Modification of NDDO approximations and application to 70 elements. Journal of Molecular Modeling, 13, 1173–1213.CrossRefGoogle Scholar
  37. Sulimov, A. V., Kutov, D. C., Katkova, E. V., Ilin, I. S., & Sulimov, V. B. (2017). New generation of docking programs: Supercomputer validation of force fields and quantum-chemical methods for docking. Journal of Molecular Graphics and Modelling, 78, 139–147.CrossRefGoogle Scholar
  38. Trott, O., & Olson, A. J. (2010). AutoDockVina: Improvising the speed and accuracy of docking with a new scoring function, efficient optimisation and multithreading. Journal of Computational Chemistry, 31, 455–461.Google Scholar
  39. Venugopala, K. N., Rashmi, V., & Odhav, B. (2013). Review on natural coumarin lead compounds for their pharmacological activity. BioMed Research International.
  40. Yadav, I. S., Nandekar, P. P., Shrivastava, S., Sangamwar, A., Chaudhury, A., & Agarwal, S. M. (2014). Ensemble docking and molecular dynamics identify knoevenagel curcumin derivatives with potent anti-EGFR activity. Gene, 539(1), 82–90.CrossRefGoogle Scholar
  41. Yeager, A. R., & Finney, N. S. (2004). The first direct evaluation of the two-active site mechanism for chitin synthase. The Journal of Organic Chemistry, 69(3), 613–618.CrossRefGoogle Scholar
  42. Young, D. C. (2009). Computational drug design: a guide for computational and medicinal chemists. John Wiley & Sons.Google Scholar

Copyright information

© Koninklijke Nederlandse Planteziektenkundige Vereniging 2018

Authors and Affiliations

  • Priscila de Andrade Gonçalves
    • 1
  • Manoelito Coelho dos Santos Junior
    • 1
  • Catiane do Sacramento Sousa
    • 2
  • Aristóteles Góes-Neto
    • 2
  • Edna Dora Martins Newman Luz
    • 3
  • Virgínia Oliveira Damaceno
    • 3
  • Ana Rosa Rocha Niella
    • 3
  • José Maria Barbosa Filho
    • 4
  • Sandra Aparecida de Assis
    • 1
    Email author
  1. 1.Departamento de SaúdeUniversidade Estadual de Feira de SantanaFeira de SantanaBrazil
  2. 2.Departamento de Ciências BiológicasUniversidade Estadual de Feira de SantanaFeira de SantanaBrazil
  3. 3.CEPLAC, Centro de Pesquisas do Cacau (CEPEC)ItabunaBrazil
  4. 4.Departamento de SaúdeUniversidade Federal da ParaíbaJoão PessoaBrazil

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