Kraft pulp bleaching and delignification by Trametes versicolor

Abstract

In 1989 we reported that the white-rot basidiomycete fungus Trametes versicolor could delignify and substantially brighten unbleached kraft pulps. Since that time, considerable effort has been dedicated to understanding the biological mechanisms of this efficient delignification system in the hope that part or all of the system can be applied industrially. Early work indicated that all components necessary for extensive kraft pulp bleaching and delignification are secreted by the fungus, but that one or more of these components must be constantly replaced or renewed. T. versicolor can bleach O 2-delignified and extended-cooking pulps as well as conventional hardwood and softwood kraft pulps. The fungus demethylates and solubilizes the residual lignin in the pulps, seemingly without depolymerization. Enzyme families secreted by T. versicolor which may be involved in delignification include lignin peroxidases (LiP), manganese peroxidases (MnP), laccases, and cellobiose dehydrogenases (CDH). Each of these families has been investigated for its interactions with kraft pulp lignin. Two laccase isozymes (laccase I and II) have been purified. The reactivities of the two isozymes on all substrates tested, including kraft lignin, were similar, with no evidence of lignin depolymerization by either enzyme. However the presence of the mediator 2,2′-azinobis (3-ethylbenzthiazoline-6-sulphonate) (ABTS) prevented and reversed the polymerization of kraft lignin by either laccase. Both laccases were able to delignify kraft pulp, but only in the presence of the mediator. Secreted MnP activity and the presence of substantial available Mn(II) ions appear necessary for delignification and pulp brightening by T. versicolor. Mutants unable to secrete MnP cannot delignify. Purified MnP substantially delignifies pulp when supplied with Mn (II), H 2O 2 and Mn (II)-complexing agents. Furthermore, the resulting pulp is made easier to bleach to high brightness with alkaline hydrogen peroxide. LiP, while produced under secondary metabolic conditions, appears to play no role in the delignification of kraft pulp by T. versicolor. White-rot fungi typically carry out net oxidation of the lignin and hydrolytic depolymerization of the carbohydrate in wood. Is there likely to be a significant role in delignification for enzymes which oxidize wood carbohydrates and reduce lignin? The family of carbohydrate-oxidizing, quinone-, metal ion- and organic free radical-reducing enzymes known as the cellobiose oxidases and cellobiose dehydrogenases (CBO, CDH, formerly CBQase) are reported to occur in a number of wood-degrading fungi. A CDH from T. versicolor has been purified, characterized and shown to decolorize and prevent laccase-mediated lignin polymerization. It can help supply MnP with chelator, hydrogen peroxide, and manganese. The rate of diffusion of enzymes or mediators in the kraft fiber wall may be a limiting factor for delignification. In order to visualize these diffusion bottlenecks we have built a model of the secondary wall and compared the pore sizes in the wall with the sizes of potential diffusible delignification agents.