Abstract:

Electrophoresis-purity peroxidase was extracted from fresh cucumber and purified through homogenization,
extraction, ammonium sulfate precipitation, CM-Sepharose and Superdex-200 chromatography. The specific activity,
recovery and purification fold of the peroxidase were 64 177.67 U/mg, 9.58% and 61.93, respectively. Molecular mass of
this purified enzyme was 40.21 kD as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE),
while native gel filtration confirmed a monomer form of 41.15 kD. Besides, the peroxidase, whose optimum temperature
and pH were 60 ℃ and 6, respectively, was comparatively stable in the temperature range of 25-45 ℃ and pH range of 5-9.
The purified peroxidase showed Km value of 53.79 mmol/L under definite conditions. In addition, its activity was scarcely
affected by potassium thiocyanate (KSCN). The peroxidase was found to be activated by urea, K+, Mn2+, Ca2+, Mg2+, Ba2+
and Cu2+ by 12%, 27%, 13%, 28%, 39%, 99% and 248% at 50 mmol/L, respectively. However, the peroxidase activity was
significantly inhibited by SDS, ascorbic acid and oxalic acid. Moreover, Zn2+, methanol, ethanol and isopropanol caused
partial inhibitory effects on the peroxidase.

Key words: cucumber, peroxidase, isolation and purification, characterization