The immobilization experiment was carried out by suspending
4 mg of CMMC into 10 ml of sodium citrate buffer (0.1 M, pH 4.0)
containing 1 mg ml1 of laccase. The resulting mixture of the supports
and laccase solution was then incubated at 25 C with shaking
at 100 rpm for 3 h to reach adsorption equilibrium. The laccase
immobilized on CMMC particles was subsequently separated magnetically,
washed with the same buffer, dried at 40 C under vacuum
using lyophilizer and stored at 4 C for the further use. To
investigate the effects of initial laccase concentration on the
adsorption capacity of the support, it was changed between 0.1
and 4.0 mg ml1. Each set of experiments was carried out in triplicate,
and the arithmetic mean values were calculated, with the
standard deviations less than 5%. The amount of laccase immobilized
on CMMC particles was calculated by determining the initial
and final concentration of laccase in liquid solution using Lowry’s
method (Lowry et al., 1951)