Purification and enzymatic properties of a textile dye-decolourizing peroxidase from Moringa oleifera roots / Purificação e propriedades enzimáticas de uma peroxidase descorante de corantes têxteis a partir de raízes de Moringa oleifera

Marina Gabrielle Guimarães de Almeida, Larissa Alves Lopes, Lucas Pinheiro Dias, Helen Paula Silva da Costa, Celso Shiniti Nagano, Rômulo Farias Carneiro, Ilka Maria Vasconcelos, Daniele de Oliveira Bezerra de Sousa


Peroxidases are ubiquitous enzymes involved in the oxidation of a variety of aromatic substrates including textile dyes, which are harmful for aquatic life and human health. Hence, the present study describes the purification of a peroxidase, named MoPOX, from Moringa oleifera roots using DEAE-Sephacel and gel filtration chromatography on a Superdex® 75 column. The peptide sequences recorded by mass spectrometry analysis confirmed the identity of MoPOX with other plant peroxidases. The optimum pH and temperature of enzyme activity were 5.2 and 70 °C, respectively. Its enzymatic activity in the presence of metal ions and classical peroxidase inhibitors was also evaluated. MoPOX follows Michaelis-Menten kinetics, with specificity, in ascending order, to the substrates ABTS < eugenol < O-dianisidine ≅ guaiacol, besides being highly thermostable. The purified peroxidase (0.015, 0.030 or 0.150 mg/mL) degraded different dyes (50 or 100 mg/L), such as Remazol® Blue RGB, Remazol® Navy RGB and Telon® Turquoise M-5G 85%. Decolorization rates varied from 15 to 90% depending on the dye concentration, enzyme concentration and exposure time. MoPOX is the first peroxidase purified from M. oleifera roots, and the results showed it has biotechnological potential for biodegradation of hazardous compounds.



Plant peroxidase, textile dye, Moringa oleifera, bioremediation.


R. V. Khandare, S.P. Govindwar, Phytoremediation of textile dyes and effluents: Current scenario and future prospects, Biotechnol. Adv. 33 (2015) 1697–1714. doi:10.1016/j.biotechadv.2015.09.003.

P.M. Dellamatrice, M.E. Silva-Stenico, L.A.B. de Moraes, M.F. Fiore, R.T.R. Monteiro, Degradation of textile dyes by cyanobacteria, Brazilian J. Microbiol. 48 (2017) 25–31. doi:10.1016/j.bjm.2016.09.012.

H. Ben Mansour, I. Houas, F. Montassar, K. Ghedira, D. Barillier, R. Mosrati, L. Chekir-Ghedira, Alteration of in vitro and acute in vivo toxicity of textile dyeing wastewater after chemical and biological remediation, Environ. Sci. Pollut. Res. 19 (2012) 2634–2643. doi:10.1007/s11356-012-0802-7.

U. Kalsoom, S.S. Ashraf, M.A. Meetani, M.A. Rauf, H.N. Bhatti, Mechanistic study of a diazo dye degradation by Soybean Peroxidase, Chem. Cent. J. 7 (2013) 1–10. doi:10.1186/1752-153X-7-93.

S. Gita, A. Hussan, T.G. Choudhury, Impact of Textile Dyes Waste on Aquatic Environments and its Treatment, Environ. Ecol. 35 (2017) 2349–2353.

Z. Yincan, L. Yan, G. Xueyong, W. Qiao, X. Xiaoping, Decolorization of Color Index Acid Orange 20 buffer solution using horseradish peroxidase immobilized on modified PAN-beads, RSC Adv. 7 (2017) 18976–18986. doi:10.1039/c7ra01698k.

T. Chiong, S.Y. Lau, Z.H. Lek, B.Y. Koh, M.K. Danquah, Enzymatic treatment of methyl orange dye in synthetic wastewater by plant-based peroxidase enzymes, J. Environ. Chem. Eng. 4 (2016) 2500–2509. doi:10.1016/j.jece.2016.04.030.

Q. Husain, Peroxidase mediated decolorization and remediation of wastewater containing industrial dyes: A review, Rev. Environ. Sci. Biotechnol. 9 (2010) 117–140. doi:10.1007/s11157-009-9184-9.

R.R. Achar, B.K. Venkatesh, H.K. Vivek, B.S. Priya, S.N. Swamy, Caralluma umbellata Peroxidase: Biochemical Characterization and Its Detoxification Potentials in Comparison with Horseradish Peroxidase, Appl. Biochem. Biotechnol. 181 (2017) 801–812. doi:10.1007/s12010-016-2250-1.

M. Bilal, T. Rasheed, H.M.N. Iqbal, Y. Yan, Peroxidases-assisted removal of environmentally-related hazardous pollutants with reference to the reaction mechanisms of industrial dyes, Sci. Total Environ. 644 (2018) 1–13. doi:10.1016/j.scitotenv.2018.06.274.

N. Šekuljica, N. Prlainović, A.B. Stefanović, M.G. Žuža, D.Z. Čičkarić, D. Mijin, Z.D. Knezevic-Jugovic, Decolorization of anthraquinonic dyes from textile effluent using horseradish peroxidase: Optimization and kinetic study, Sci. World J. 2015 (2015). doi:10.1155/2015/371625.

B.S. Janović, A.R. Collins, Z.M. Vujčić, M.T. Vujčić, Acidic horseradish peroxidase activity abolishes genotoxicity of common dyes, J. Hazard. Mater. 321 (2017) 576–585. doi:10.1016/j.jhazmat.2016.09.037.

A.I. Maizuwo, A.S. Hassan, H. Momoh, J.A. Muhammad, Phytochemical constituents, biological activities, therapeutic potentials and nutritional values of Moringa oleifera (Zogale): A review, J. Drug Des. Med. Chem. 3 (2017) 60–66. doi:10.11648/j.jddmc.20170304.12.

D.R. Hoagland, D.I. Arnon, The water-culture method for growing plants without soil, Calif. Agr. Expt. Sta. Circ. 347 (1950) 1–32. doi:citeulike-article-id:9455435.

U.K. Laemmli, Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature. 227 (1970) 680–685. doi:10.1038/228549a0.

H. Urbanek, K. Kuzniak-Gebarowska, E. Herka, Elicitation of defence responses in bean leaves by Botrytis cinerea polygalacturonase, Acta Physiol. Plant. 13 (1991) 43–50.

P.K. Smith, R.I. Krohn, G.T. Hermanson, A.K. Mallia, F.H. Gartner, M.D. Provenzano, E.K. Fujimoto, N.M. Goeke, B.J. Olson, D.C. Klenk, Measurement of protein using bicinchoninic acid, Anal. Biochem. 150 (1985) 76–85. doi:10.1016/0003-2697(85)90442-7.

M.M. Bradford, A Rapid and sensitive method for the quantitation microgram quantities of protein utilizing the principle of protein-dye binding reproduction, 72 (1976) 248–254. doi:10.1016/0003-2697(76)90527-3.

G. Candiano, M. Bruschi, L. Musante, L. Santucci, G.M. Ghiggeri, B. Carnemolla, P. Orecchia, L. Zardi, P.G. Righetti, Blue silver: a very sensitive colloidal Coomassie G-250 staining for proteome analysis, Electrophoresis. 25 (2004) 1327–1333. doi:10.1002/elps.200305844.

A. Shevchenko, H. Tomas, J. Havliš, J. V. Olsen, M. Mann, In-gel digestion for mass spectrometric characterization of proteins and proteomes, Nat. Protoc. 1 (2007) 2856–2860. doi:10.1038/nprot.2006.468.

N. Fawal, Q. Li, B. Savelli, M. Brette, G. Passaia, M. Fabre, C. Mathé, C. Dunand, PeroxiBase: a database for large-scale evolutionary analysis of peroxidases, Nucleic Acids Res. 41 (2013) 441–444. doi:10.1093/nar/gks1083.

R.M. Zacharius, T.E. Zell, J.H. Morrison, J.J. Woodlock, Glycoprotein staining following electrophoresis on acrylamide gels, Anal Biochem. 30 (1969) 148–152. doi:10.1016/0003-2697(69)90383-2.

M. Dubois, K.A. Gilles, J.K. Hamilton, P.A. Rebers, F. Smith, Calorimetric method for determination of sugars and related substances, Anal. Chem. 28 (1956) 350–356. doi:10.1021/ac60111a017.

S. Johri, U. Jamwal, S. Rasool, A. Kumar, V. Verma, G.N. Qazi, Purification and characterization of peroxidases from Withania somnifera (AGB 002) and their ability to oxidize IAA, Plant Sci. 169 (2005) 1014–1021. doi:10.1016/j.plantsci.2005.05.015.

S. Khatun, Ashraduzzaman, R. Karim, F. Pervin, N. Absar, A. Rosma, Purification and characterization of peroxidase from Moringa oleifera L. leaves, BioResources. 7 (2012) 3237–3251. doi:10.15376/biores.7.3.3237-3251.

A. Altay, T. Koktepe, L. Durmaz, F. Topal, İ. Gülçin, E. Köksal, Purification and selected biochemical properties of peroxidase from cress (Lepidium sativum sub sp. sativum), Int. J. Food Prop. 21 (2018) 2610–2621. doi:10.1080/10942912.2018.1540989.

S.A. Basha, U.J.S. Prasada Rao, Purification and characterization of peroxidase from sprouted green gram (Vigna radiata) roots and removal of phenol and p-chlorophenol by immobilized peroxidase, J. Sci. Food Agric. 97 (2017) 3249–3260. doi:10.1002/jsfa.8173.

O. Márquez, K.N. Waliszewski, R.M. Oliart, V.T. Pardio, Purification and characterization of cell wall-bound peroxidase from vanilla bean, LWT - Food Sci. Technol. 41 (2008) 1372–1379. doi:10.1016/j.lwt.2007.08.017.

M.K. Das, R.S. Sharma, V. Mishra, A novel cationic peroxidase (VanPrx) from a hemi-parasitic plant (Viscum angulatum) of Western Ghats (India): Purification, characterization and kinetic properties, J. Mol. Catal. B Enzym. 71 (2011) 63–70. doi:10.1016/j.molcatb.2011.03.010.

C. Theivarasu, S. Venkatachalam, Purification and characterization of membrane-bound Borassus flabellifer L. thermostable peroxidase, Indian Jounal Biochem. Biophys. 55 (2018) 273–279.

D.A. Converso, M.E. Fernandez, Peroxidase isozymes from wheat germ: purification and properties, Phytochemistry. 40 (1995) 1341–1345.

A.M. Elsayed, U.M. Hegazy, M.G.A. Hegazy, S.S. Abdel-Ghany, W.H. Salama, A.M.H. Salem, A.S. Fahmy, Purification and biochemical characterization of peroxidase isoenzymes from Ficus carica latex, Biocatal. Agric. Biotechnol. 16 (2018) 1–9. doi:10.1016/j.bcab.2018.07.009.

M. Movahedi, S.Z.A. Samsam Shariat, H. Nazem, Immobilization of Lactoperoxidase on Graphene Oxide Nanosheets and Copper Oxide Nanoparticles and Evaluation of Their Stability, Catal. Letters. 149 (2018). doi:10.1007/s10562-018-2620-0.

P.K. Robinson, Enzymes: principles and biotechnological applications, Essays Biochem. 59 (2015) 1–41. doi:10.1042/bse0590001.

Y.M. Tao, S. Wang, H.L. Luo, W.W. Yan, Peroxidase from jackfruit: Purification, characterization and thermal inactivation, Int. J. Biol. Macromol. 114 (2018) 898–905. doi:10.1016/j.ijbiomac.2018.04.007.

N. V. Di Russo, D.A. Estrin, M.A. Martí, A.E. Roitberg, pH-Dependent Conformational Changes in Proteins and Their Effect on Experimental pKas: The Case of Nitrophorin 4, PLoS Comput. Biol. 8 (2012). doi:10.1371/journal.pcbi.1002761.

M. Shahnawaz Khan, M. Saud Al-Bagmi, M. Alhasan Ismael, A.M. Al-Senaidy, A. Ben Bacha, F. Mabood Husain, An efficient methodology for the purification of date palm peroxidase: Stability comparison with horseradish peroxidase (HRP), Saudi J. Biol. Sci. (2018). doi:10.1016/j.sjbs.2018.04.002.

U. Kalsoom, H.N. Bhatti, M. Asgher, Characterization of Plant Peroxidases and Their Potential for Degradation of Dyes: a Review, Appl. Biochem. Biotechnol. 176 (2015) 1529–1550. doi:10.1007/s12010-015-1674-3.

B. Sharma, A.K. Dangi, P. Shukla, Contemporary enzyme based technologies for bioremediation: A review, J. Environ. Manage. 210 (2018) 10–22. doi:10.1016/j.jenvman.2017.12.075.

V.P. Pandey, U.N. Dwivedi, Purification and characterization of peroxidase from Leucaena leucocephala, a tree legume, J. Mol. Catal. B Enzym. 68 (2011) 168–173. doi:10.1007/s12010-014-0776-7.

I.O. Adewale, A.T. Adekunle, Biochemical properties of peroxidase from white and red cultivars of kolanut (Cola nitida), Biocatal. Agric. Biotechnol. 14 (2018) 1–9. doi:10.1016/j.bcab.2018.01.013.

H.P. Oliveira, R.G.G. Silva, J.T.A. Oliveira, D.O.B. Sousa, M.L. Pereira, P.F.N. Souza, A.A. Soares, V.M. Gomes, A.C.O. Monteiro-Moreira, F.B.M.B. Moreno, I.M. Vasconcelos, A novel peroxidase purified from Marsdenia megalantha latex inhibits phytopathogenic fungi mediated by cell membrane permeabilization, Int. J. Biol. Macromol. 96 (2017) 743–753. doi:10.1016/j.ijbiomac.2016.12.083.

J.O. Rojas-Reyes, V. Robles-Olvera, O. Carvajal-Zarrabal, C. Castro Matinez, K.N. Waliszewski, M.G. Aguilar-Uscanga, Purification and characterization of peroxidase from avocado (Persea americana mill, cv. hass), J. Sci. Food Agric. 94 (2014) 1844–1853. doi:10.1002/jsfa.6503.

E. Dubrovskaya, N. Pozdnyakova, S. Golubev, A. Muratova, V. Grinev, A. Bondarenkova, O. Turkovskaya, Peroxidases from root exudates of Medicago sativa and Sorghum bicolor: Catalytic properties and involvement in PAH degradation, Chemosphere. 169 (2017) 224–232. doi:10.1016/j.chemosphere.2016.11.027.

Y. Anita, A. Sundowo, N.L.P. Dewi, E. Filailla, H. Mulyani, C. Risdian, S. Banjarnahor, M. Hanafi, E.P. Istyastono, Biotransformation of Eugenol to Dehydroeugenol Catalyzed by Brassica juncea Peroxidase and its Cytotoxicity Activities, Procedia Chem. 16 (2015) 265–271. doi:10.1016/j.proche.2015.12.049.

O.M. Darwesh, I.A. Matter, M.F. Eida, Development of Peroxidase Enzyme Immobilized Magnetic Nanoparticles for Bioremediation of Textile Wastewater Dye, J. Environ. Chem. Eng. (2018). doi:10.1016/j.jece.2018.11.049.

A.H. Alneyadi, I. Shah, S.F. Abuqamar, S.S. Ashraf, Differential degradation and detoxification of an aromatic pollutant by two different peroxidases, Biomolecules. 7 (2017) 1–18. doi:10.3390/biom7010031.

J.D. Baumer, A. Valério, S.M.A.G.U. Souza, G.S. Erzinger, A. Furigo, A.A.U. Souza, Toxicity of enzymatically decolored textile dyes solution by horseradish peroxidase, J. Hazard. Mater. 360 (2018) 82–88. doi:10.1016/j.jhazmat.2018.07.102.

S.M.A.G. Ulson de Souza, E. Forgiarini, A.A. Ulson de Souza, Toxicity of textile dyes and their degradation by the enzyme horseradish peroxidase (HRP), J. Hazard. Mater. 147 (2007) 1073–1078. doi:10.1016/j.jhazmat.2007.06.003.

T. Marchis, P. Avetta, A. Bianco-Prevot, D. Fabbri, G. Viscardi, E. Laurenti, Oxidative degradation of Remazol Turquoise Blue G 133 by soybean peroxidase, J. Inorg. Biochem. 105 (2011) 321–327. doi:10.1016/j.jinorgbio.2010.11.009.

G. Arabaci, A. Usluoglu, The enzymatic decolorization of textile dyes by the immobilized polyphenol oxidase from quince leaves, Sci. World J. 2014 (2014). doi:10.1155/2014/685975.

DOI: https://doi.org/10.34117/bjdv6n4-066


  • There are currently no refbacks.