Elsevier

Organic Geochemistry

Volume 33, Issue 2, February 2002, Pages 111-124
Organic Geochemistry

Lignin demethylation and polysaccharide decomposition in spruce sapwood degraded by brown rot fungi

https://doi.org/10.1016/S0146-6380(01)00144-9Get rights and content

Abstract

The organic residues produced in the brown-rot (BR) of wood by many basidiomycetes fungi are ubiquitous on most coniferous forest floors. This degraded wood tissue is characterized by low levels of polysaccharides and a high proportion of demethylated lignin with minor glycerol side chain oxidation. Because of the selective enrichment in an aromatic dihydroxy-rich lignin residue, the chemical and biological reactivity of BR degraded wood will be distinctly different from white rot, the other primary class of fungal wood decay, which typically produces oxidized, lignin-depleted residues. The biochemical mechanism by which BR fungi perform this distinctive degradative chemistry is only starting to become known, and molecular studies which examine the chemical changes imparted to lignin over the long-term decay process are lacking. Using 13C-labeled tetramethylammonium hydroxide thermochemolysis (13C-TMAH) and solid state 13C NMR, we investigated the relationship between lignin oxidation/demethylation and polysaccharide metabolism in a 32-week time series study of spruce sapwood inoculated with either of two BR fungi (Postia placenta and Gloeophyllum trabeum). Our findings demonstrate a close relationship between lignin demethylation and polysaccharide loss and suggest demethylation may play a mechanistic role in polysaccharide loss, possibly by assisting in Fenton reactions where catechol/quinone oxidation and cycling aids in iron reduction. The residue remaining after 16 weeks of decay is devoid of polysaccharides, in contrast to the 68% polysaccharide carbon present in the initial spruce, and exhibits an increased aromatic dihydroxy content (resulting from demethylation of the 3-methoxyl carbon) of up to 22% of the lignin, as determined by 13C-TMAH thermochemolysis. In a typical soil or porewater environment these chemical changes would make BR residues highly reactive toward redox sensitive polyvalent metals (e.g. ferric iron) and likely to adsorb to metal hydroxide surfaces.

Introduction

Wood decay fungi are essential for sustained productivity in terrestrial ecosystems. In addition to their role in metal and nutrient cycling in aerobic environments (Jellison et al., 1997, Ostrofsky et al., 1997, Connolly et al., 1999, Hobbie et al., 1999), they are the main biological agents for the conversion of vascular plant tissue to humic materials (e.g. Blanchette et al., 1994, Eriksson et al., 1990) which are ultimately mineralized by soil fungi and bacteria. On a global basis, vascular plant tissue makes up the largest living pool of actively cycling organic matter on earth. Lignin, a primarily ether-linked phenylpropanoid biopolymer found in wood cells and in small amounts in foliar tissue, makes up the second most abundant biopolymer after cellulose. Brown-rot (BR) basidiomycete fungi, the dominant wood decay fungi in coniferous forests, play a unique role in the conversion of wood to coarse debris and soil organic matter, as they are selective for the metabolism of the polysaccharide components of wood over that of lignin (e.g. Blanchette et al., 1994) (see Fig. 1). White-rot fungi, in contrast, can degrade lignin and cellulose simultaneously or selectively degrade lignin (Eriksson et al., 1990). During BR degradation, the methoxyl carbons of lignin are removed, generating an aromatic hydroxyl-rich product abundant in ortho-hydroxy substitution (Ander et al., 1988, Agusin et al., 1989, Enoki et al., 1988, Jin et al., 1990, Filley et al., 2000). Further lignin alteration is thought to be quantitatively minimal but associated with oxidation of the aliphatic side chains and potentially limited lignin depolymerization (e.g. Agosin et al., 1989).

The mechanism of BR decay is not yet determined, but is the focus of active research in mycology, plant pathology and soil sciences. The initial stages of decay are thought to involve the action of Fenton chemistry (Fe2++H2O2) for the production of hydroxyl anions and radicals (Koenigs, 1974). Phenolic compounds produced by the fungi function as ferric iron chelators and sources of electrons for iron reduction (e.g. Chanhoke et al., 1991, Enoki et al., 1997, Goodell et al., 1997, Kerem et al., 1999, Paszczynski et al., 1999). These low molecular weight reactants, unlike enzymes, are small enough to penetrate the sound wood lignocellulose fabric, and have been shown in immunolabeling studies to be present throughout the S2 layer of the BR-degraded cell wall (Jellison et al., 1997). Additionally, many studies have demonstrated that the lignin-rich residues remaining after BR decay can be enriched in metals, particularly calcium, iron, manganese, and magnesium (Ostrofsky et al., 1997, Jellison et al., 1997). BR residues, therefore, are potential sites for enhanced metal concentration and sources of aromatic dihydroxy-rich, altered lignin to forest soils and groundwater.

The extent to which lignin derived carbon is partitioned in soil and water and influences the biogeochemistry of its surroundings is controlled by many factors including its original chemical and structural composition and, in no small part, the nature of the chemical alteration imparted to it during microbial degradation (e.g. Bianchi et al., 1997, Hedges and Oades, 1997). For example, the chemical functionality of degraded plant components (e.g. polyphenolic aromatic acids and alcohols or aliphatic acids) determines the ability of soil and dissolved organic matter to chelate and retain metals such as iron, aluminum, calcium and magnesium (e.g. Tan and Binger, 1986, Schnitzer et al., 1984, Xu and Goodell, 2001). Also, the presence of phenolic compounds, in particular ortho-hydroxy aromatic alcohols (catechols) enables facile redox reactions with ferric iron (e.g. Voelker and Sulzberger, 1996, Paszczynski et al., 1999). The binding capacity of soil and dissolved organic matter for cations and mineral surfaces significantly influences residence time and preservation of organic matter pools with different functional groups (e.g. Miltner and Zech, 1998, Schmidt et al., 2000). The ability of organic matter to effectively bind to metal hydroxy oxides, or clay surfaces bears directly on the efficiency of sorptive protection mechanisms in soils (Henrichs, 1995, Kaiser and Guggenberger, 2000).

We present here the results from a study of spruce sapwood degraded by two common BR basidiomycete fungi, Gloeophyllum trabeum and Postia placenta, in a 32-week laboratory time series inoculation study. The overall goal of this study was to access the chemical alterations imparted to lignin and cellulose during this little understood but ubiquitous process, and address any interdependent relationships in the chemistry of degradation of these two biopolymers by BR fungi. We conducted a molecular biomarker (using 13C-tetramethylammonium hydroxide thermochemolysis) and bulk spectroscopic (using solid-state 13C NMR) assessment of fresh and degraded residues. Herein, we discuss the relationships between lignin demethylation (catechol production), wood mass loss, and polysaccharide (cellulose and hemicellulose) depletion in BR wood and relate the identified chemistry to potential geochemical influence in soils.

Section snippets

Brown-rot inoculation of spruce sapwood

Red spruce (Picea rubens (Sarg.)) sapwood blocks were degraded using a modified ASTM soil block procedure (ASTM, 1994). The soil mix (80 g) was added to 500 ml glass chambers along with 200 ml distilled and deionized water. Two birch (Betula spp.) feeder strips (1×40×40 mm) were placed on the soil surface in each jar and inoculated with four small (2×2 mm) blocks of malt agar supporting the growth of either Postia placenta (Mad-698-R) or Gloeophyllum trabeum (Mad-617-R). Spruce blocks (40×20×20

Weight loss during decay

With increasing time in the degradation experiment the wood blocks changed in color, shape, and weight in a fashion characteristic of BR decay. The blocks progressively turned dark brown taking on a friable texture and shrinking in volume with time. The changes were first visible at 2 weeks time near the base of the blocks in contact with the inoculation strip. By 4 weeks the base of the blocks had shrunk significantly and the originally rectangular blocks took on an inverted trapezoidal shape

Comparison of P. placenta and G. trabeum-induced wood alteration

The lignin demethylation and polysaccharide loss exhibited in G. trabeum and P. placenta degradation (Fig. 5, Fig. 6) is consistent with the known utilization pathways of BR fungi (e.g. Enoki et al., 1988, Jin et al., 1990, Blanchette et al., 1994, Filley et al., 2000). Important insights into BR chemistry and possibly lignin structure are revealed by closer examination of the differences in the relative degree of demethylation among the monomers and in the relative amount of polysaccharide and

Conclusion

The 13C-TMAH thermochemolysis of the residues remaining after a 32 month inoculation experiment using two BR fungi, G. trabeum and P. placenta, showed progressive demethylation of all monomers with a mean demethylation at 22%. G. trabeum degraded wood exhibited a greater degree of demethylation compared to P. Placenta degraded wood. Wood decayed by P. placenta exhibited a greater degree of lignin side chain oxidation as determined by yield of G6. The demethylation of the lignin macromolecule

Acknowledgments

The authors would like to acknowledge helpful comments regarding this manuscript by Rose Filley and Mark Teece and two additional reviewers. We would also like to acknowledge the support of USDA grants 97-34158-502301-35103-09935 and the Carnegie Institution of Washington. This is paper 2520 of the Maine Agricultural and Forestry Experiment Station.

Associate EditorJ. Rice

References (46)

  • P.G. Hatcher et al.

    Comparison of two thermochemolytic methods for the analysis of lignin in decomposing gymnosperm woodthe CuO oxidation method and the method of thermochemolysis with tetramethylammonium hydroxide (TMAH)

    Organic Geochemistry

    (1995)
  • P.G. Hatcher et al.

    Comment on the origin of benzenecarboxylic acids in pyrolysis methylation studies

    Organic Geochemistry

    (1995)
  • P.G. Hatcher et al.

    Comparison of dehydrogenase polymer (DHP) lignin with native lignin from gymnosperm wood by thermochemolysis using tetramethylammonium hydroxide (TMAH)

    Organic Geochemistry

    (1996)
  • J.I. Hedges et al.

    Comparative organic geochemistries of soils and marine sediments

    Organic Geochemistry

    (1997)
  • S.M. Henrichs

    Sedimentary organic matter preservationan assessment and speculative synthesis—a comment

    Marine Chemistry

    (1995)
  • J. Jellison et al.

    The role of cations in the biodegradation of wood by the brown rot fungi

    International Biodeterioration and Biodegradation

    (1997)
  • Z. Kerem et al.

    Biodegradative mechanism of the brown rot basidiomycete Gloeophyllum trabeumevidence for an extracellular hydroquinone-driven fenton reaction

    Federation of European Biochemical Societies

    (1999)
  • L.M. Mayer

    Relationships between mineral surfaces and organic carbon concentrations in soils and sediments

    Chemical Geology

    (1994)
  • A. Miltner et al.

    Beech leaf litter lignin degradation and transformation as influenced by mineral phases

    Organic Geochemistry

    (1998)
  • M. Schnitzer et al.

    Kinetics and characteristics of humic acids produce from simple phenols

    Soil Biol. Biochem.

    (1984)
  • American Society for Testing and Materials, 1994. Standard method of accelerated laboratory test of natural decay...
  • P. Ander et al.

    Cleavage and metabolism of methoxyl groups from vanillic and ferulic acids by brown-rot and soft-rot fungi

    Cellulose Chemical Technology

    (1988)
  • R.A. Blanchette et al.

    Biodegradation of compression wood and tension wood by white and brown rot fungi

    Holzforschung

    (1994)
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