Continuous biotransformation of pyrogallol to purpurogallin using cross-linked enzyme crystals of laccase as catalyst in a packed-bed reactor

Abstract

Cross-linked enzyme crystals (CLEC) of laccase were prepared by crystallizing laccase with 75% (NH4)(2)SO4 and cross-linking using 1.5% glutaraldehyde. The cross-linked enzyme crystals were further coated with 1 mmol L-1 beta-cyclodextrin by lyophilization. The lyophilized enzyme crystals were used as such for the biotransformation of pyrogallol to purpurogallin in a packed-bed reactor. The maximum conversion (76.28%) was obtained with 3 mmol L-1 pyrogallol at a residence time of 7.1 s. The maximum productivity (269.03 g L-1 h(-1)) of purpurogallin was obtained with 5 mmol L-1 pyrogallol at a residence time of 3.5 s. The productivity was found to be 261.14 g L-1 h(-1) and 251.1 g L-1 h(-1) when concentrations of 3 mmol L-1 and 7 mmol L-1 respectively were used. The reaction rate of purpurogallin synthesis was maximum (2241.94 mg purpurogallin mg(-1) CLEC h(-1)) at a residence time of 3.5 s, when 5 mmol L` pyrogallol was used as the substrate. The catalyst to product ratio calculated for the present biotransformation was 1:2241. The CLEC laccase had very high stability in reuse and even after 650 h of continuous use, the enzyme did not lose its activity.