Investigations on the laccase-catalyzed polymerization of lignin model compounds using size-exclusion HPLC

The reaction of laccase from Trametes hirsuta with monomeric and dimeric lignin model compounds was studied with special attention to the formation of polymeric reaction products. Different HPLC techniques were evaluated for the analysis of the reaction products. Methods requiring a solvent change from aqueous to organic phase were found not to be adequate for the analysis of product mixtures since evaporation over nitrogen led to the polymerization of reactive monomers and extraction with dichloromethane was not quantitative in all cases. Therefore, aqueous phase size-exclusion chromatography (SEC) at high pH was used to characterize reaction product mixtures. The product molecular weight distribution (MWD) after reaction with laccase was observed to be highly dependent on the type of model compound whereas the enzyme concentration had only a limited impact.