Covalent immobilizing horseradish peroxidase on electrochemically-functionalized biochar for phenol removal

Enzyme-based biocatalytic treatment has been known as an effective measure to biologically degrade organic pollutants. Advantageously, enzymes could be immobilized on solid supports, and such fact enables reuse/prolong the enzymatic capability. It could be of great importance to functionalize a support material for enhancing the immobilization efficiency/stability of enzymes. As such, this study laid great emphasis on covalent bonding to immobilize horseradish peroxidase (HRP) on a functionalized rice straw biochar with glutaraldehyde (GA) as a crosslinker. Biochar was pretreated by the electrochemical method and the acid treatment respectively to enrich the oxygen-containing functional groups. These led to the enhanced immobilizing ability of biochar. The HRP immobilized on the electrochemically-functionalized biochar (HRP-EBC) showed three times as much enzyme activity as the HRP directly adsorbed onto biochar. The HRP immobilized on the acid-functionalized biochar (HRP-ABC) showed activity similar to that of HRP-EBC. It was concluded that both the (acid/electrochemical) pretreatments are effective to enhance enzyme immobilization. Nevertheless, the electrochemical functionalized method of biochar is chemical oxidant-free, and one important lesson from a series of tests was that the pretreatment of biochar through the electrochemical method could be more environmentally benign. Moreover, employing HRP-EBC could be beneficial from a perspective of a real environmental practice considering its higher pH, thermal stability, and good reusability. 80% of phenol was degraded in 1 h in the presence of HRP-EBC when pH was 7.0 and a ratio of H₂O₂ to phenol was 1:1.5.