Journal of
Medicinal Plants Research

  • Abbreviation: J. Med. Plants Res.
  • Language: English
  • ISSN: 1996-0875
  • DOI: 10.5897/JMPR
  • Start Year: 2007
  • Published Articles: 3605

Full Length Research Paper

Purification and characterization of peroxidase from Turkish black radish (Raphanus sativus L.)

Melda Şişecioğlu1, İlhami Gülçin1, 2*, Murat Çankaya3, Ali Atasever4, M. Hilal Şehitoğlu1, Habibe Budak Kaya1 and Hasan Özdemir1
1Department of Chemistry, Faculty of Sciences, Atatürk University, 25240-Erzurum, Turkey. 2School of Health Services, Ibrahim Çeçen University, TR-4100-Agri-Turkey. 3Department of Biology, Faculty of Sciences, Erzincan University, 24100 Erzincan, Turkey. 4Department of Food Sciences, Ispir H. Polat Vocational School, Atatürk University, 25900 Ispir, Erzurum, Turkey.
Email: [email protected], [email protected]

  •  Accepted: 02 June 2010
  •  Published: 18 June 2010

Abstract

Peroxidases (EC 1.11.1.7; donor: hydrogen peroxide oxidoreductase) are part of a large group of enzymes associated with cell wall biosynthesis, response to injury, disease, resistance and wound repair. They catalyze the oxidation of various electron donor substrates such as phenols and aromatic amines in the presence of hydrogen peroxide. In the present study, peroxidase, a primer antioxidant enzyme, was purified 9.7 fold from Turkish black radish (Raphanus sativus L.) in a yield of 2% by ammonium sulphate precipitation, dialysis and CM-Sephadex ion exchange chromatography. To check the enzyme purity, sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed and a single band was observed. The substrate specificity of peroxidase was investigated using guaiacol (2-methoxyphenol). Optimum pH, optimum temperature, optimum ionic strength, stable pH, stable temperature and thermal inactivation conditions were determined for guaiacol / H2O2substrate pattern. These kinetic properties were found to be 6.0, 30°C, 1.0 M, 9.0, and 60°C, respectively. The molecular weight (Mw) of the enzyme was found to be 66 kDa by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS - PAGE) method. Native polyacrylamide gel electrophoresis (Native-PAGE) was performed for isoenzyme determination and a single band was observed. Km and Vmax values were calculated from Lineweaver-Burk graphs.

 

Key words: Turkish black radish, Raphanus sativus, peroxidase, enzyme purification, enzyme characterization.

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