Abstract:

Mixtures of nanogold sol(NGS) and multi-walled carbon nanotubes(MWCNTs) were used as a carrier for laccase immobilization. Three immobilization approaches were evaluated via comparing the amount of immobilized laccase and their specific activity of immobilized enzyme. Thermal stability and reusability of immobilized laccase were investigated and the effects of particle size of nanogold sol particles on laccase immobilization and immobilized enzyme kinetics were examined. Results from experiments reveal that NGS/MWCNTs matrices have the most excellent capability of laccase immobilization and high specific activity of immobilized laccase. Amount of immobilized enzyme and specific activity reach 33.80 mg/g and 9.433 U/mg, respectively, through simple physical adsorption when NGS particle size is 37 nm. It is also shown that the strategy of physical adsorption is suitable for immobilization of laccase in practice according to comprehensive evaluation of the amount of immobilized laccase, thermal stability, specific activity and reusability. Immobilized laccase on NGS/MWCNTs through covalent coupling can retain about 75％of initial activity after 2 h at 70 ℃ and retain 70％of initial activity after 20 recycles of repeated use. Conclusion from experiments indicates smaller NGS particle(24 nm) makes the affinity between immobilized laccase and its substrate(KM=0.027 mmol/L) stronger and the apparent catalyzing rate constant(1088 min-1) greater.

Key words: Laccase, enzyme immobilization, nano-gold sol, particle size effects, thermal stability, reusability