Abstract: Conventional pulp-bleaching techniques with chlorine or chlorine-based chemicals can, under certain conditions, generate chlorinated organic compounds that are toxic to the environment. The pulp and paper industry is facing an increasing pressure from environmentally concerned organizations to replace the conventional bleaching techniques with environmentally benign ones. Enzymatic bleaching methods have recently drawn much attention as being environmentally friendly. In addition to xylanase, laccase has been the most actively investigated enzyme for biobleaching of kraft pulp because laccase can be produced in large amounts at a reasonable price and use cheap oxygen as an electron acceptor. However, expensive redox mediators are still a hurdle in the implementation of laccase in pulp bleaching.
Laccase (EC 1.10.3.1) belongs to a family of multi-copper oxidases that are widespread in numerous fungi, in various plant species (18), in the bacterium Azospirillum lipoferum (10), and in a dozen of studied insects (25). Laccase has various functions, including participation in lignin biosynthesis (21), plant pathogenicity (22), the degradation of plant cell walls (12, 17), insect sclerotization (3), bacterial melanization (10), and melanin-related virulence for humans (26). Chemically, all of these functions of laccases are related to oxidation of a range of aromatic substances. However, the net effect of such oxidations could be very different and even work in opposite directions. Plant laccases, for example, oxidize monolignols to form polymeric lignins, whereas laccases from white-rot fungi degrade and depolymerize lignins.
In the degradation of lignin by white-rot fungi, the redox potential of the lignin-degrading enzymes has long been believed to play a crucial role because nonphenolic subunits, the most predominant lignin substructures in wood, have high redox potentials. The well-studied lignin peroxidase is able to oxidize nonphenolic aromatic compounds with very high ionization potentials such as 1,2-dimethoxybenzene (E1/2 = 1,500 mV) and veratryl alcohol (14, 20). Lignin peroxidase was thus once believed to be a key enzyme for fungal degradation of lignin, whereas laccase was believed to be less important because it could not oxidize veratryl alcohol (a typical model compound for nonphenolic lignin). The highest redox potential of a laccase reported so far does not exceed 800 mV, which is believed not to be high enough to oxidize a nonphenolic lignin structure. However, it has been demonstrated that laccase is able to oxidize some compounds (redox mediators) with a higher redox potential than laccase itself, although the mechanism by which this happens is not known (2, 7). In the presence of such redox mediators, laccase is also able to oxidize nonphenolic lignin model compounds and decrease pulp kappa number to a great extent (5, 8). Several effective redox mediators have been reported so far (2, 5, 6, 8, 13). The importance of the redox potential of laccases in the oxidation of lignin model compounds by laccase/mediator systems will be addressed here.
While much effort has been devoted to search for more effective redox mediators, the laccase parameters governing lignin degradation and pulp bleaching are still not fully elucidated. In an effort to determine these parameters, we compared the ability of different laccases for the oxidation of lignin model compounds in a laccase-mediator system. More specifically, four laccases from different fungal species were purified and used to oxidize the β-O-4 dimer I (the most predominant lignin substructure) and phenol red (a phenolic lignin model compound). Laccases from the different sources were found to oxidize dimer I and phenol red at different rates. Criteria for a better laccase and more effective laccase-mediator systems for pulp bleaching have been suggested.