TRANSFORMATION OF LIGNIN-RELATED COMPOUNDS WITH LACCASE IN ORGANIC-SOLVENTS

The extracellular laccase (benzenediol: oxygen oxidoreductase, EC 1.10 .3.2) of Trametes versicolor was isolated from culture medium and immo bilized by entrapment of the enzyme in a solvent-resistant hydrophilic matrix like Sepharose-CL-6B. The gel-enzyme association has been show n to be stable in water containing organic solvents. The efficiency of the immobilized laccase in different organic solvents was comparable with the activity shown in a buffered aqueous system. The immobilized laccase in organic solvents showed a good stability and a high toleran ce to elevated temperatures. Water-insoluble organosolv lignin (OL), d issolved in dioxane/water, was readily converted by immobilized laccas e from Trametes versicolor. The transformed lignin showed an increase in phenolic groups, changes in the quantity of conjugated elements, an d a significant modification of both the aliphatic and aromatic carbon moieties of the lignin molecule. The changes in the lignin molecule w ere analyzed by UV-, IR-spectroscopy, and C-13-NMR solid-state spectro scopy. High-performance size exclusion chromatography (HPSEC) of the l ignin transformed with the laccase-Sepharose complex revealed a pronou nced increase in weight-average molecular weight. Polymerization of th e lignin in the organic solvent proved to be 4 times more effective th an polymerization of the same compound in an aqueous system. Water-ins oluble organosolv lignin as well as a variety of lignin-related aromat ics, solubilized in dioxane-H2O (7:3), was readily converted by laccas e preparation either in batch or in continuous flow column. Reaction o f laccase with the solubilized lignin generates in the reaction media reduced oxygen species able to reduce the cytochrome c. For the first time it is now possible to perform enzymatically catalyzed reactions w ith lignin in an organic solvent. This is a first step towards an enzy matically derivatization of lignin, the formation of polymer blends on the basis of lignin by an enzymatically catalyzed reaction.