Removal of Anthraquinone Dye by Cross-Linked Enzyme Aggregates From Fresh Horseradish Extract

Horseradish roots (Armoracia rusticana) fresh extract was the source of unpurified horseradish peroxidase (HRP) which was used for the immobilization in a form of cross-linked enzyme aggregates (CLEAs). Among three precipitants used, acetone appeared to be the best choice for the precipitation. Acetone with 2% of glutaraldehyde gave the recovery of 30.3% of the enzyme initial activity. Horseradish peroxidase immobilized in a form of cross-linked enzyme aggregates (HRP-CLEAs) showed remarkable efficiency in Acid Violet 109 (AV 109) oxidation. Under optimal conditions (pH 4, dye concentration 30 mg L-1, hydrogen peroxide concentration 0.1 mM), 72.4 and 88.9% of dye was oxidized using HRP-CLEAs in a batch and packed bed reactor, respectively. Higher stability was apparent in the packed bed reactor compared to the batch reactor where the mechanical instability of aggregates under constant agitation has been proved. After seven consecutive cycles the retained activity was 60 and 20% in the recycled packed bed and batch reactor, respectively. The feasibility of HRP-CLEAs application in dye oxidation was confirmed by significant dye oxidation percentage and reduction of the toxicity in the samples collected after enzymatic treatment in the recycled packed bed reactor, using brine shrimp, Artemia salina.