Keywords
proteases
karatasin
2D zymography
How to Cite
##article.highlights##
- The proteolytic activity of B. karatas (8.59 U·mg-1) was greater than that of Ananas comosus (3.42 U·mg-1).
- The activity was higher at pH 6 and 7 and in concentrations lower than 5 % NaCl.
- Proteases have thermal stability at 30 and 50 °C for 210 minutes.
- The fruits of B. karatas are potentially useful to obtain proteases for industrial use.
Abstract
Introduction: Bromeliads are a source of proteases that have not been fully studied. Objectives: To evaluate the proteolytic activity of wild fruits of Bromelia karatas L. under different temperature, pH and NaCl conditions, and to estimate the thermal stability of their proteases. Materials and methods: The effect of pH (6, 7, 8, 9, 10 and 12), incubation temperatures (30, 40, 50, 60 and 70 °C) and NaCl concentrations (5, 10 and 20 %) on the proteolytic activity of the fruits was compared. The thermal stability of the proteases was assessed at 30, 50 and 70 °C for 240 minutes. The molecular weights and isoelectric points of the proteases were estimated by non-reducing two-dimensional zymography. The data were analyzed using a one-factor ANOVA and Tukey's test (α = 0.05). Results and discussion: The proteolytic activity of B. karatas fruit extract (8.59 U·mg-1) was greater than that of Ananas comosus [L.] Merr. variety comosus (3.42 U·mg-1). The activity was higher at pH 6 and 7 and in concentrations lower than 5 % NaCl. Proteases were stable at 30 and 50 °C for 210 minutes. The two-dimensional zymogram, under non-reducing conditions, showed at least 40 light zones with apparent molecular weights between 27.3 and 290 kDa, potentially representing proteases. Conclusion: The proteases of B. karatas have potential for application in the food industry.References
Arshad, Z. I. M., Amid, A., Yusof, F., Jaswir, I., Ahmad, K., & Loke, S. P. (2014). Bromelain: overview of industrial application and purification strategies. Applied Microbiology and Biotechnology, 98(17), 7283–7297. doi: https://doi.org/10.1007/s00253-014-5889-y
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248–254. Retrieved from http://hoffman.cm.utexas.edu/courses/bradford_assay.pdf
Bruno, M. A., Pardo, M. F., Caffini, N. O., & López, L. M. I. (2002). Purification of a new endopeptidase isolated from fruits of Bromelia hieronymi Mez (Bromeliaceae). Acta Farmaceutica Bonaerense, 21(1), 51–56. Retrieved from https://www.researchgate.net/publication/267974610_Purification_of_a_New_Endopeptidase_Isolated_from_Fruits_of_Bromelia_hieronymi_Mez_Bromeliaceae
Daniel, R. M., Dines, M., & Petach, H. H. (1996). The denaturation and degradation of stable enzymes at high temperatures. Biochemical Journal, 317, 1-11. Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1217448/pdf/8694749.pdf
de Lencastre, N. L. C., Jozala, A. F., Lopes, A. M., Santos-Ebinuma, C. V, Mazzola, P. G., & Pessoa, J. A. (2016). Stability, purification, and applications of bromelain: A review. Biotechnology Progress, 32(1), 5–13. doi: https://doi.org/10.1002/btpr.2190
Espejo-Serna, A., & López-Ferrari, A. R. (2010). Flora del Bajío y de regiones adyacentes: Bromeliaceae. Retrieved from http://www1.inecol.edu.mx/publicaciones/resumeness/FLOBA/Bromeliacaeae 165(1).pdf
Espejo-Serna, A., López-Ferrari, A. R., & Ramírez-Morillo, I. (2005). Bromeliaceae. Flora de Veracruz, 136, 1–103. Retrieved from http://www1.inecol.edu.mx/publicaciones/resumeness/FLOVER/136-Espejo_I.pdf
González-Salvatierra, C., Andrade, J. L., Orellana, R., Peña-Rodríguez, L. M., & Reyes-García, C. (2013). Microambiente lumínico y morfología y fisiología foliar de Bromelia karatas (Bromeliaceae) en una selva baja caducifolia de Yucatán, México. Botanical Sciences, 91(1), 75–84. Retrieved from http://www.scielo.org.mx/scielo.php?pid=S2007-42982013000100006&script=sci_arttext
Guadix, A., Guadix, E. M., Paéz-Dueñas, M. P., González-Tello, P., & Camacho, F. (2000). Procesos tecnológicos y métodos de control en la hidrólisis de proteínas. Ars Pharmaceutica, 41(1), 79–89. Retrieved from http://revistaseug.ugr.es/index.php/ars/article/view/5735
Kozlowski, L. P. (2016). IPC – Isoelectric point calculator. Biology Direct, 11, 55. doi: https://doi.org/10.1186/s13062-016-0159-9
Larocca, M., Rossano, R., Santamaria, M., & Riccio, P. (2010). Analysis of pineapple [Ananas comosus (L.) Merr.] fruit proteinases by 2-D zymography and direct identification of the major zymographic spots by mass spectrometry. Food Chemistry, 123(4), 1334–1342. doi: https://doi.org/10.1016/j.foodchem.2010.06.016
Li, Y., Yu, J., Goktepe, I., & Ahmedna, M. (2016). The potential of papain and alcalase enzymes and process optimizations to reduce allergenic gliadins in wheat flour. Food Chemistry, 196(1), 1338–1345. doi: https://doi.org/10.1016/j.foodchem.2015.10.089
Meza-Espinoza, L., Vivar-Vera, M. A., García-Magaña, M. L., Sáyago-Ayerdi, S., Chacón-López, A., Becerraea-Verdín, E., & Montalvo-González, E. (2017). Enzyme activity and partial characterization of proteases obtained from Bromelia karatas fruit and compared with Bromelia pinguin proteases. Food Science and Biotechnology, 27(2), 509–517. doi: https://doi.org/10.1007/s10068-017-0244-6
Miranda, F. (1958). Estudios acerca de la vegetación. In E. Beltrán (Ed.), Los recursos naturales del sureste y su aprovechamiento (pp. 215–271). México: Instituto Mexicano de Recursos Naturales Renovables, A. C.
Monteiro, R. F., & Forzza, R. C. (2016). Typification of Bromelia karatas, the type of the genus Bromelia (Bromeliaceae). Taxon, 65(5), 1101–1106. doi: https://doi.org/10.12705/655.12
Montes, C., Amador, M., Cuevas, D., & Córdoba, F. (1990). Subunit structure of karatasin, the proteinase isolated from Bromelia plumieri (karatas). Agricultural and Biological Chemistry, 54(1), 17–24. Retrieved from https://www.jstage.jst.go.jp/article/bbb1961/54/1/54_1_17/_pdf
Montes, R. C., Terán, G. V. F., Zuñiga, B. R. A., & Caldón, Y. E. (2014). Descripción morfológica de Bromelia karatas, recurso genético promisorio. Biotecnología en el Sector Agropecuario y Agroindustrial, 12(1), 62–70. Retrieved from http://www.scielo.org.co/pdf/bsaa/v12n1/v12n1a08.pdf
Moreno-Hernández, J. M., Hernández-Mancillas, X. D., Navarrete, E. L. C., Mazorra-Manzano, M. A., Osuna-Ruiz, I., Rodríguez-Tirado, V. A., & Salazar-Leyva, J. A. (2017). Partial characterization of the proteolytic properties of an enzymatic extract from “Aguama” Bromelia pinguin L. fruit grown in Mexico. Applied Biochemistry and Biotechnology, 182(1), 181–196. doi: https://doi.org/10.1007/s12010-016-2319-x
Moyano, D. D., Osorio, R. M., Murillo, P. E., Murillo, A. W., Solanilla, D. J., Méndez, A. J., & Aristizabal, S. J. (2012). Evaluación de parámetros bromatológicos, fitoquímicos y funcionalidad antioxidante de frutos de Bromelia karatas (Bromeliaceae). Vitae, 19(1), S439–S441. Retrieved from http://www.redalyc.org/pdf/1698/169823914138.pdf
Natalucci, C. L., Brullo, A., López, L. M. I., Hilal, R. M., & Caffini, N. O. (1996). Macrodontin, a new protease isolated from fruits of Pseudananas macrodontes (Morr) Harms (Bromeliaceae). Journal of Food Biochemistry, 19(6), 443–454. doi: https://doi.org/10.1111/j.1745-4514.1995.tb00547.x
Natalucci, C. L., Payrol, J. A., Bruno, M. A., Martin, M. I., Pardo, M. F., Pérez, A. T., …Caffini, N. O. (2009). Fuentes naturales de fitoproteasas: proteasas de Bromeliaceae. In Ciencia y Tecnología para el Desarrollo – CYTED (Ed.), Enzimas proteolíticas de vegetales superiores. Aplicaciones industriales (pp. 41–50). Buenos Aires, Argentina: Mariscal & Asociados.
Payrol, J. A., Obregón, W. D., Trejo, S. A., & Caffini, N. O. (2008). Purification and characterization of four new cysteine endopeptidases from fruits of Bromelia pinguin L. grown in Cuba. The Protein Journal, 27(2), 88–96. doi: https://doi.org/10.1007/s10930-007-9111-2
Peña, F., R., Duran, O. D. S., & Baleta, M. L. C. (2015). Efecto del marinado con NaCl y tripolifosfato-sódico sobre las propiedades bromatológicas en carne de cabra. Biotecnología en el Sector Agropecuario y Agroindustrial, 13(1), 64–72. Retrieved from http://www.scielo.org.co/pdf/bsaa/v13n1/v13n1a08.pdf
R Core Team (2017). R: A language and environment for statistical computing. Vienna, Austria: R Foundation for Statistical Computing.
Ramli, A. N., Aznan, T. N., & Illias, R. M. (2017). Bromelain: from production to commercialization. Journal of the Science of Food Agricultural, 97(5), 1386–1395. doi: https://doi.org/10.1002/jsfa.8122
Zar, J. H. (1999). Biostatistical analysis (4th ed.). New Jersey: Prentice Hall.
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