Jerzy Trojanowski

Decomposition of 14C-labelled lignin, holocellulose and lignocellulose by mycorrhizal fungi

Five different species of known ecto-mycorrhizal fungi: Cenococcum geophilum, Amanita muscaria, Tricholoma aurantium, Rhizopogon luteolus and Rhizopogon roseolus were studied for their ability to metabolize the major components of plant cell walls. All strains were able to decompose 14C-labelled plant lignin, 14C-lignocellulose and 14C-DHP-lignin at a rate which was lower than the one observed for the known white rot fungi Heterobasidion annosum and Sporotrichum pulverulentum. Also 14C-(U)-holocellulose was relatively less degradable for the mycorrhizal fungi than for the white rotters. On the other hand, aromatic monomers like 14C-vanillic acid were decomposed to a much higher extent by two species of mycorrhizal fungi compared to the activity observed for Heterobasidion annosum. The results of the experiments reveal that these stains of mycorrhizal fungi are well able to utilize the major components of plant material and thus can contribute to litter decomposition in the forest floor.

Laccase activities of Penicillium chrysogenum in relation to lignin degradation

Abstract An extracellular laccase capable of oxidizing ABTS (the diammonium salt of 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid) was detected in ligninolytic cultures of Penicillium chrysogenum. By contrast, no lignin peroxidase, manganese-dependent peroxidase or aryl-alcohol oxidase was detected at any time during culturing. Both ABTS laccase activity and mineralization of dehydrogenative polymerizate of coniferyl alcohol were regulated by the C/N ratio in the medium and partially inhibited in the presence of thioglycolic acid, suggesting that both events are associated. In the presence of several known laccase inducers neither ABTS laccase activity nor mineralization rates were enhanced. However, a new laccase was detected in P. chrysogenum, able to oxidize 2,6-dimethoxyphenol but not involved in lignin mineralization. Studies with the known ligninolytic basidiomycete Trametes villosa suggest that lignin degradation by this fungus also involves the action of laccase.

The Effects of Fungal Peroxidase on Na-Lignosulfonates

Peroxidase (EC 1.11.1.7) from Trametes versicolor fungus was separated from laccase (EC 1.10.3.2) by the affmity chromatography method. The purified peroxidase was immobilized on glass beads using glutaraldehyde äs a spacer arm. Three fractions of Na-lignosulfonates isolated from the cellulose industry wastes were incubated with the immobilized fungal peroxidase and H2O2 from 3 to 24 hours. Using a Sephadex G-50 gel chromatography column it was established that the molecular weight of Na-lignosulfonates changed after the time of incubation. The peroxidase action caused both polymerization and depolymerization of Na-lignosulfonate molecuies. The molecular weight of Na-lignosulfonates was determined before and after the immobilized fungal peroxidase treatment.

Mikrobieller Abbau von 14C- und 32S-markierten Ligninsulfonsäuren durch Pilze, Bakterien und Mischkulturen

Zusammenfassung Coder S-markierte Ligninsulfonsäuren (LS) wurden durch Umsetzung von spezifisch C-markiertem DHP-Lignin aus Coniferylalkohol oder im Ligninanteil markiertem Pflanzenmaterial (OCH3, 0-C oder C-Ring) mit Na-Bisulfit oder durch Umsetzung unmarkierten Lignins mit S-markiertem Na-Bisulfit hergestellt. Die C-markierten LS-Präparate wurden mit Weißoder Moderfäulepilzen oder Bakterien inkubiert und die CO2-Freisetzung gemessen. Im Vergleich zum natürlichen Lignin war diese Freisetzung wesentlich geringer. Durch eine vorhergehende UV-Bestrahlung konnte die CO2Freisetzung gesteigert werden. Einige Bakterien, besonders Pseudomonaden, bewirkten eine beträchtliche Abspaltung der Sulfonatgruppe als Sulfat. Wurde die C-LS vorher einige Tage mit Pseudomonaden und anschließend mit Weißoder Moderfäulepilzen inkubiert, so war die CO2-Freisetzung stark erhöht.

Studies of enzymatic oxidation of TMP-fibers and lignin model compounds by a Laccase–Mediator-System using different 14C and 13C techniques

In this work, the results of the enzymatic oxidation of TMP-fibers (thermomechanical pulp) and a well-structured lignin model compound, the dehydropolymer (DHP), were investigated by different 14C and 13C methods, caused by a Laccase–Mediator-System (LMS). These methods are the nuclear magnetic resonance spectroscopy (13C-NMR) with DHP (unmarked) and the determination of the 14CO2 release of 14C-marked DHP and TMP-fibers. The 13C-NMR measurements were chosen to analyze the structural changes of the LMS-treated DHP model compounds and TMP-fibers qualitatively and quantitatively. The data of 14CO2 release give an explanation of the demethylation of DHP and TMP-fibers. The effect of the LMS is shown by comparing the results in respect of DHP and TMP-fibers, which were only treated with laccase and of an inactivated LMS as the control. Comparing the results of the 13C-NMR method, in particular the use of the Mediator during the enzymatical treatment, showed significant changes in the structure of the DHP. Also, the TMP-fibers were materially influenced by the LMS. The analysis of the 14CO2 release data of the 14C-marked DHP and TMP-fibers revealed that the rate of 14CO2 increases in the 14C-2 atom as well as in the O14CH3 group within the first hour of Laccase–Mediator incubation. Therefore, the 14CO2 release from the DHP was higher than from the TMP-fibers.

New form of lignolytically active mycelium generated by immobilization of protoplasts isolated from the white rot fungi Heterobasidion annosum and Polyporus pinsitus

SummaryImmobilized mycelia regenerated from immobilized protoplasts isolated from lignin-degrading Basiodiomycetes have been shown to be able to decompose specifically 14C-labelled dehydropolymers of coniferylalcohol (DHP-lignin) and monomeric lignin-related compounds more intensively than native mycelium, by decarboxylation, demethylation, ring and side chain cleavage. Protoplasts of two white rot fungi were immobilized by entrapment in Na- alginate gel and remained intact after the immobilization procedure. Within the first 3 days of incubation in culture medium, regeneration of hyphal cells occurred. Since hyphal cells regenerated from protoplasts within gel beads were hindered from stretching by the matrix, the microbial immobilized cells differed from native mycelium in terms of their morphology. The time course and extent of lignin degradation by native mycelium and regenerated mycelium of the examined white rot fungi also differed, a sign that there may also be differences between them in terms of the physiology of lignin degradation.

DEGRADATION OF LIGNIN AND LIGNIN RELATED-COMPOUNDS BY PROTOPLASTS ISOLATED FROM FOMES-ANNOSUS

Protoplasts from a lignolytic fungus Fomes annosus were prepared through enzymatic hydrolysis of mycelium utilizing Novozym, a wall lytic enzyme preparation. Isolated protoplasts and living mycelium were compared in their ability to degrade 14C-labelled lignin related phenols and dehydropolymers of labelled coniferyl alcohol (synthetic lignin). The amounts of 14CO2 released from O14CH3-groups, 14C-2′-side chains and 14C-rings by protoplasts was in the same range as those for intact mycelium. The methoxyl groups of synthetic lignin were more rapidly metabolized by protoplasts than by mycelium. When calculated in dpm of released 14CO2 per mg protein the decomposition of 14C-labelled synthetic lignin and lignin-related monomers in a hyphae-free system of protoplasts was considerable higher than that obtained by the intact mycelium. The presence of intact hyphae is thus not necessary for lignin degradation to occur.

Laccase-Mediator Catalyzed Modification of Wood Fibers: Studies on the Reaction Mechanism and Making of Medium-Density Fiberboard

ABSTRACT Owing to the constant increase of prices of the process for petrochemical resin and the possibility of harmful formaldehyde emissions from industrial produced medium-density fiberboards (M...