Catalytic and kinetic properties of purified Eichhornia crassipes leaf peroxidase

Abstract

Peroxidase from Eichhornia crassipes leaf was purified 23.58 fold with 18.58% yield by means of (NH 4 ) 2 SO 4 precipitation, ion exchange and Sephadex G-75 gel filtration chromatography. Optimum temperature and substrate-dependent pH optimum for enzyme activity were 40 °C and pH 4.0, 9.0 and 6.0 for 2,2'-azino-bis-(3-ethylbenzthiazolin)-6-sulfonate (ABTS), guaiacol and pyrogallol, respectively. The enzyme had high pH stability and moderate thermal stability at temperatures up to 60 °C; the activation energy of inactivation of the enzyme was ~122.29 kJ mol -1 . Temperature-denaturing and spontaneous recovery were shown to be time-dependent while Ca 2+ -enhanced recovery of the denatured enzyme was in a time-dependent manner. The enzyme showed preferential affinity for ABTS over guaiacol and pyrogallol with K M values of 31.11, 21.91 and 6.45 mM respectively. It was reversibly inhibited by Pb 2+ , Hg 2+ and EDTA while urea only caused loss of ~30.40% activity after 60 min of incubation. E. crassipes leaf peroxidase has potential use for broad range of applications.

Stability profile of <i>Pistia stratiotes</i> leaf peroxidase

“…The optimum pH for P. stratiotes leaf peroxidase was 6.0 with the optimum range between 5.0 and 7.0. This is in agreement with previous studies where peroxidases from different sources have been reported to have pH optima often within the range 4.5-6.5 (Tomas-Barberan et al, 1997;Bestwick et al, 1998;Thongsook and Barrett, 2005;Onsa et al, 2006;Hu et al, 2012;Khatun et al, 2012;Arise et al, 2016). The shift in optimum pH from 6.0 to 6.5, however, indicate that Pistia sratiotes leaf peroxidase largely retains its enzymatic activity over a narrow range of pH after 60 minutes of incubation.…”

“…The optimum temperature of 40ºC obtained was similar to that reported by other authors from different sources (Altunkaya and Gokmen, 2011;Hu et al, 2012;Arise et al, 2016). Bhatti et al (2006) reported that lettuce stem peroxidase has an optimum temperature of 45˚C, and peroxidase from cauliflower had optimum temperature of 30ºC (Koksal and Gulcin (2008).…”

“…Peroxidases from plant sources are being studied extensively due to their broad substrate specificity, distributions and potential industrial applications (Carlos et al, 2004;Aehle, 2007;Hamid and Khalil-ur-Rehman, 2009;Arise et al, 2016). In this study, purification of Pistia stratiotes leaf peroxidase was achieved through a threestep procedure that included crude extraction, ammonium sulphate precipitation and gel filtration.…”

“…They are heme-containing enzymes which catalyse the single one electron oxidation of several substrates at the expense of H2O2 (Galati and O'Brien, 2004;Veitch, 2004). Due to their broad substrate specificity, peroxidases have importance in industrial applications such as decolorization of dyes, removal of phenolic contaminants, and production of analytic and diagnostic kits, ELISA, biosensors, and pharmaceuticals (Clarke et al, 1993;Ruzgas et al, 1996;Agostini et al, 2002;Carlos et al, 2004;Aehle, 2007;Hamid and Khalil-ur-Rehman, 2009;Ng et al, 2010;Arise et al, 2016).…”

“…Recently, Horseradish peroxidase (HRP) is being exploited for cancer treatment (Veitch et al, 2004, Arise et al, 2016Bonifert et al, 2016). HRP is the most extensively studied plant peroxidase due to its wide applicability (Regalado et al, 2004).…”