Biochemical Engineering Journal 46 (2009) 96–104 Contents lists available at ScienceDirect Biochemical Engineering Journal journal homepage: www.elsevier.com/locate/bej Applications of Celite-adsorbed white radish (Raphanus sativus) peroxidase in batch process and continuous reactor for the degradation of reactive dyes Rukhsana Satar, Qayyum Husain ∗ Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh 202002, UP, India article info Article history: Received 17 January 2009 Received in revised form 8 April 2009 Accepted 11 April 2009 Keywords: Celite Decolorization Reactive dyes Immobilized Batch process White radish peroxidase abstract White radish peroxidase immobilized on Celite has been employed for the treatment of reactive dyes: Reactive Red 120 and Reactive Blue 171. A comparative study of decolorization of these dyes by soluble and immobilized white radish peroxidase was made. Effect of different redox mediators: 1- hydroxybenzotriazole, syringaldehyde, verataryl alcohol and violuric acid on the decolorization of reactive dyes by white radish peroxidase was tested. The investigated dyes were decolorized to different extents in the presence of these mediators. However, 1-hydroxybenzotriazole was found to be the most effec- tive mediator for decolorization of Reactive Red 120 and Reactive Blue 171 by white radish peroxidase. Maximum decolorization of the dyes was observed at pH 5.0 and 40 ◦ C in 1 h. Comparative operational stability performance of soluble and immobilized white radish peroxidase for the treatment of dyes was checked in the presence of various denaturing and inhibiting agents such as sodium azide, organic sol- vents and mercuric chloride. Among the studied dyes, white radish peroxidase exhibited significantly higher affinity for Reactive Red 120. Toxicity of the dyes was tested by Allium cepa test. Immobilized per- oxidase decolorized dyes more effectively in batch process. Absorption spectra of treated and untreated dye solutions were found to show a marked difference. Efficiency of immobilized peroxidase was checked in a continuous reactor where the immobilized enzyme exhibited 73% decolorization of Reactive Red 120 even after 1 month of operation of the reactor. © 2009 Elsevier B.V. All rights reserved. 1. Introduction Enzyme based procedures have attracted attention of the researchers for targeting aromatic pollutants as a potential and viable alternative to conventional methods due to their highly selective nature. The inhibition by toxic substances in enzymatic treatment is minimum and the process can be operated over a broad range of aromatic concentrations with low retention time [1]. Peroxidases are versatile catalysts with wide spectrum appli- cations in a number of industrial processes and have already been employed for the transformation of toxic compounds of industrial origin such as aromatic compounds to preserve the quality of water [1–3]. Soluble enzymes exhibit some inherent limitations such as reusability, stability and use in continuous reactors [4,5]. Such lim- itations could be avoided by using enzymes in their immobilized form that can act as catalysts with long lifetime and high stabil- Abbreviations: DMF, dimethylformamide; DMSO, dimethylsulfoxide; WRP, white radish peroxidase; S-WRP, soluble WRP; I-WRP, immobilized WRP. ∗ Corresponding author. Tel.: +91 571 2700741; fax: +91 571 2706002. E-mail addresses: qayyumhusain@yahoo.co.in, qayyumbio@rediffmail.com (Q. Husain). ity [6,7], thus being a better alternative for wastewater treatment at large scale. Immobilization protects enzymes from denaturation and helps to retain them in biochemical reactors to further cat- alyze the subsequent feed and offer more economical exploitation of biocatalysts in industry, waste treatment and in the develop- ment of bioprocess monitoring devices like the biosensor [8–11]. Immobilization helps in maintaining homogeneity of enzymes in the reaction media since it avoids aggregation of enzyme particles. It is an efficient way to prevent inactivation and extend enzyme half-life. Immobilization procedures can also increase structural rigidity of the enzyme thus improving pH, temperature and/or organic solvent tolerance [12,13]. In addition to the ease of han- dling, immobilized enzymes are well suited for use in continuous reactors. Since the loss of activity is a certainty over time, the sup- port material should be easy to be regenerated with active enzyme [14,15]. Several methods have been employed for enzyme immo- bilization, which include adsorption onto insoluble materials, entrapment in polymeric gels, encapsulation in membranes, chem- ical crosslinking by using bifunctional or multifunctional reagents and linking to an insoluble carrier. Adsorption is a very simple, cheap and effective but frequently reversible process [7,9]. Celite is an impervious, chemically inert and solid support widely employed for the adsorption or deposition of biocatalysts [7,15]. Diatomite car- 1369-703X/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.bej.2009.04.012