6.1 Introduction
The peroxidase isolated from horseradish, HRP, is the most available and commonly used peroxidase. One factor that has limited its widespread and large-scale use is its high cost of production. A cost effective purification technology and exploringalternative sources with high peroxidase activity can help to bring down the cost of enzyme production. Peroxidase from roots of Raphanus sativus can serve as a cost effective alternative for HRP.
Gil-Rodríguez et al. (2008) purified the peroxidase from Japanese radish by several purification steps. The juice extracted from roots of radish was ultra-centrifuged using 10 kDa membrane. The retentate was then loaded on Macro-Prep High-S medium at pH 6.1 and eluted with a linear gradient of 0–1 M NaCl. The eluate was further loaded on Macro-Prep High-Q medium
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UF systems commonly consist of three steps:
(1) A pre-concentration step achieving similar concentrations of low molecular weight components in retentate and permeate
(2) A diafiltration step to purify retentate by addition of diafiltration liquid, and
(3) A final concentration step to maximize the concentration of high molecular weight solutes in retentate. Literature on purification of peroxidase from Raphanus sativus is santand requires minimum number of purification steps to minimize the cost. The current study focuses on developing a purification process for radish peroxidase from the roots of Raphanus sativus.
Since plant peroxidases are present in low concentration in the extract, its recovery and purification involves traditional downstream processing steps. The grinding/extraction, precipitation/concentration and different chromatographic separation techniques are the general steps involved in purification of enzymes (Fig. 6.1). Figure 6.1 Purification protocol for peroxidase from Raphanus