Extraction, purification and characterization of peroxidase from Vitis vinifera wastes

Document Type: Research Paper


1 Department of Biology, Faculty of Sciences, Payame Noor University, Tehran, Iran

2 Department of Biology, University of Guilan, Rasht, Iran


The proper consumption of plant wastes could not only provide a possible source of natural products, but it also is an environmental friendly research. The aim of this study was to use grape wastes as a source of peroxidase.  In practice, one isoenzyme of peroxidase (POD1) was partially purified from Vitis vinifera wastes, the plant which is widely harvested in Iran. The activity of this novel peroxidase was determined using guaiacol as its substrate. The new peroxidase was partially purified and its kinetic parameters determined. The values of Km and Vmax of peroxidase for guaiacol were 83.2 mM and 0.35 M/min respectively. Optimum pH and temperature were determined for guaiacol to be 6.2 and 60°C respectively. According to SDS-PAGE results, the molecular weight of isozymes was 38-40 KD. The results indicate that agricultural leftovers from Vitis vinifera are a considerable source for a peroxidase with reliable kinetic behaviors.


Abbaspour, N & Babaee, L 2017, Effect of salicylic acid application on oxidative damage and antioxidant activity of grape (Vitis vinifera L.) under drought stress condition, International Journal of Horticultural Science and Technology, 4: 29-50.

Acharya, AP, Nafisi, PM, Gardner, A, MacKay, JL, Kundu, K, Kumar, S & Murthy, N 2013, "A fluorescent peroxidase probe increases the sensitivity of commercial ELISAs by two orders of magnitude". Chemical Communications, 49: 10379-10381.

Adam, W, Lazarus, M, Saha-Moller, CR, Weichold, O, Hoch, U & Schreier P 1999, Bio-transformations with peroxidases, Advances in Biochemical Engineering. 63: 74-108.

Afsharnezhad, M, Shahangian, SS, Panahi, E & Sariri, R 2017, Evaluation of the antioxidant activity of extracts from some fruit peels. Caspian Journal of Environmental Sciences, 15: 213-222.

Agostini, E, Hernandez-Ruiz, J, Arnao, MB 2002, “Ascota M A peroxidase isoenzyme secreted by turnip (Brassica napus) hairy-root culture inactivation by hydrogen peroxide and application in diagnosis kits”. Journal of Biotechnology and Applied Biochemistry, 35: 1-7.

Alizadeh, Ketek Lahijani, H, Naderi Beni, A & Tavakoli, V 2018, Heavy metals in coastal sediments of South Caspian Sea: natural or anthropogenic source?Caspian Journal of Environmental Sciences, 16: 45-61.

Almagro, L, Gómez Ros, LV, Belchi-Navarro, S,  Bru, R,  Ros Barceló, A & Pedreño MA, 2008, "Class III peroxidases in plant defence reactions," Journal of Experimental Botany, 60: 377-390.

Apel, K & Hirt, H 2004, Reactive oxygen species: metabolism, oxidative stress, and signal transduction. Annual Reviews in Plant Biology, 55: 373-99.

Badan-Ara, Marzdashti, R, Aghamaali, MR, Varasteh, A, Nowruzfashkhami, MR & Sabkara F 2018, Purification and characterization of lysozyme in Persian sturgeon, Acipenser persicus (Borodin, 1897) from the Southwest Caspian Sea. Caspian Journal of Environmental Sciences 16: 359-367.

Bansal, N, Kumari L & Kanwar SS 2012, Peroxidase and its applications. International Journal of Institutional Pharmacy and Life Sciences, 2: 2249-6807.

Barceló, AR, Ros, LVG, Gabaldon, C, Serrano ML, Pomar, F, Carrion JS & Pedreno MA 2004, "Basic peroxidases: The gateway for lignin evolution?" Phytochemistry Reviews, 3: 61-78.

Battistuzzi, G, Bellei, M, Bortolotti, CA, & Sola, M 2010, Redox properties of heme peroxidases. Archives of Biochemistry and Biophysics, 500: 21-36.

Bernards, MA, Summerhurst, DK & Razem, FA 2004, "Oxidases, peroxidases and hydrogen peroxide: The suberin connection". Phytochemistry Reviews, 3: 113-126.

Bradford, MM 1976, A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye-binding. Annals of Biochemistry, 72: 248-258.

BRENDA: The Comprehensive Enzyme Information System @ Institute of Biochemistry and Bioinformatics at the Technical University of Braunschweig, Germany.

Cardinali, A, Tursi, N, Ligorio, A, Giuffrida MG, Napolitano, L, Caliandro, R, Sergio L, Di Venere D, Lattanzio V & Sonnante G 2011, Purification, biochemical characterization and cloning of a new cationic peroxidase isoenzyme from artichoke. Plant Physiology and Biochemistry, 49: 395-403.

Circu, ML & Aw, TY 2010, "Reactive oxygen species, cellular redox systems, and apoptosis". Free Radical Biology and Medicine, 48: 749-762.

Conesa, A, Punt, PJ, Cees, AMJJ & van den Hondel 2002, Fungal peroxidases: molecular aspects and applications. Journal of Biotechnology, 93: 143-158.

Cosio, C & Dunand, C 2009, Specific functions of individual class III peroxidase genes. Journal of Experimental Botany, 60: 391-408.

Dąbrowska, G, Kata, A, Goc, A, Szechyńska-Hebda, M & Skrzypek E 2007, Characteristics of the plant ascorbate peroxidase family. Acta Biologica Cracoviensia Series Botanica, 49: 7-17.

Guo, XH, Lv YD, Jiang J, Li HY & Liu GF 2012, Expression, purification and characterization of a peroxidase from Tamarix hispida. African Journal of Biotechnology, 11: 1540-1544.

Hancock, JT, Desikan, R & Neill SJ 2001, Role of reactive oxygen species in cell signaling pathways. Biochemical and Biomedical Aspects of Oxidative Modification, 29: 345-350.

He F 2011, Laemmli-SDS-PAGE,Bio-protocolBio101: e80.

Heuzé, V, Thiollet, H & Tran G 2017, Grape leaves and vine shoots. Feedipedia, a programme by INRA, CIRAD, AFZ and FAO.

Khan, AA & Robinson, DS 1993, Purification of an anionic peroxidase isoenzyme from mango (Mangifera indica L. var. Chaunsa). Food Chemistry, 46: 61-64.

Köksal, E & Gülçin, I 2008, Purification and Characterization of Peroxidase from Cauliflower (Brassica oleracea L. var. botrytis) Buds, Protein & Peptide Letters, 15: 320-326.

Kvaratskhelia, M, Winkel, C & Thorneley, RNF 1997, Purification and characterization of a nove1 class III peroxidase isoenzyme from tea leaves. Plant Physiology, 114: 1237-1245.

Liu, HL, Van, X, Hong X & Kong, LY 2007, Biotransformation of sinapic acid catalyzed by Momordica charantia peroxidase. Journal of Agricultural and Food Chemistry, 55: 1003-1008.

Mafulul, SG, Joel, EB, Barde, LA, Lenka, JL,  Ameh, AA & Phililus, MG 2018, Extraction, partial purification and characterization of peroxidase from Calotropis procera leaves. Journal of Advances in Biology & Biotechnology, 18: 1-10.

Mathé, C, Barre, A, Jourda C & Dunand, C 2010, Evolution and expression of class III peroxidases. Archives of Biochemistry and Biophysics, 500: 58-65.

Mazaheri, M & Fergusen, J 2018, Bacterial responses to environmental herbicide pollutants (glyphosate and paraquat).Caspian Journal of Environmental Sciences, 16: 35-43.

Mujeeb, F, Khan, AF, Bajpai, P & Pathak N (2018), Profiling of enzymatic antioxidants in leaf extracts of different bael varieties/accessions. Research Journal of Pharmaceutical, Biological and Chemical Sciences, 9: 1483-1498.

Murakami, S, Takahara, H & Shiraiwa M 2007, Purification and characterization of three neutral extracellular isoperoxidases from rye leaves. Phytochemistry, 68: 777-784.

O’Brien, PJ 2000, Peroxidases, Chemico-Biological Interactions, 129: 113-139.

Regalado, G, Perez-Arvizu, O, Garcia-Almendarez, B & Whitaker, JR 1999, Purification and properties of two acid peroxidases from Brussels sprouts. Journal of Food Biochemistry, 23: 435-450.

Saboora, A, Parsiavash, L & Moosavi-Nejad, Z 2012, Purification and kinetic properties of guaiacol peroxidase in turnip (Brassica napus var. okapi) root during different growth stages. Progress in Biological Sciences, 2: 76-86.

Sakharov, IY, Vesga, MK, Galaev, IY, Sakharova, IV & Pletjushkina, OY 2001, Peroxidase from leaves of royal palm tree Roystonea regia: purification and some properties. Plant Science, 161: 853–860.

Saraiva, JA, Nunes, CS & Coimbra, MA 2007, Purification and characterization of olive (Olea europaea L.) peroxidase – Evidence for the occurrence of a pectin binding peroxidase. Food Chemistry, 101: 1571-1579.

Şişecioğlu, M, Gülçin, I, Çankaya, M, Atasever, A, Şehitoğlu, MH, Budak, Kaya H & Özdemir, H 2010, Purification and characterization of peroxidase from Turkish black radish (Raphanus sativus L.). Journal of Medicinal Plants Research, 4: 1187-1196.

Vamos-Vigyazo, L 1981, Polyphenol oxidase and peroxidase in fruits and vegetables. CRC Critical Reviews in Food Science and Nutrition, 15: 49-127.

Wall, SB, Oh, JY, Diers AR & Landar, A 2012, Oxidative modification of proteins: An emerging mechanism of cell signaling. Frontiers in Physiology, 3: 369-378.

Watanabe, L, Nascimento, LS,  Zamorano, LS, Shnyrov, VL & Polikarpov, I 2007, Purification, crystallization and preliminary X-ray diffraction analysis of royal palm tree (Roystonea regia) peroxidase. Acta Crystallography Section F Structural Biology Crystalization Communication, 63: 780-783.

Welinder, KG, Gajhede, M 1993, in: Welinder KG, Rasmussen SK, Penel C & Greppin H (Eds.), Plant peroxidases: Biochemistry and Physiology, University of Geneva, 35-42.

Zámocký, M, Hofbauer, S, Schaffner I, Gasselhuber, B, Nicolussi, A, Soudi M,  Paul, G, Furtmüller KF & Obinger, C 2015, Independent evolution of four heme peroxidase superfamilies, Archives of Biochemistry and Biophysics, 574: 108-119.

Zhang, JQ, Li  GP, Kang YL, Teng, BH & Yao 2017, Biomimetic synthesis of resveratrol trimers catalyzed by horseradish peroxidase. Molecules, 22: 819-825.