Solid-state fermentation,lignin degradation and resulting digestibility of wheat straw fermented by selected white-rot fungi

Summary Lignin biodegradation, carbon loss and in vitro dry matter digestibility (IVDMD) have been investigated during the solid state fermentation of wheat straw by eight previously selected strains of white-rot fungi. A mathematical model of the degradation kinetics is presented. [The time period required to reach maximum rates of ¹⁴CO₂ and unlabeled CO₂ release from (¹⁴C)-lignin-labelled wheat straw and from whole wheat straw, respectively, was generally short (6–10 days).] High rates of ¹⁴C-lignin degradation were achieved by Pycnoporus cinnabarinus (2.9% ¹⁴CO₂ evolved/day), an unidentified strain Nancon (3.0%/day), Sporotrichum pulverulentum Nov. (3.4%/day), Bjerkandera adusta (2.4%/day), and Dichomitus squalens (2.3%). However, only the latter two strains degraded whole wheat straw slowly and Bjerkandera adusta was not able to degrade more than 23% of the ¹⁴C-lignin. Cyathus stercoreus and Dichomitus squalens facilitated the highest improvement in IVDMD (68% against 38% for the sound straw) after 20 and 15 days of cultivation respectively, with low dry matter losses (15–20%). A study of the fate of ¹⁴C-lignin during fermentation using these two fungal strains showed that maximal levels of (¹⁴C)-water-soluble compounds are reached before peak levels of ¹⁴CO₂ evolution suggesting that these compounds are intermediates in lignin degradation. A possible relationship between water-soluble lignins and IVDMD improvement is discussed.     

Potential of selected fungal species to degrade wheat straw,the most abundant plant raw material in Europe

Background

Structural component of plant biomass, lignocellulose, is the most abundant renewable resource in nature. Lignin is the most recalcitrant natural aromatic polymer and its degradation presents great challenge. Nowadays, the special attention is given to biological delignification, the process where white-rot fungi take the crucial place owing to strong ligninolytic enzyme system. However, fungal species, even strains, differ in potential to produce high active ligninolytic enzymes and consequently to delignify plant biomass. Therefore, the goals of the study were characterization of Mn-oxidizing peroxidases and laccases of numerous mushrooms as well as determination of their potential to delignify wheat straw, the plant raw material that, according to annual yield, takes the first place in Europe and the second one in the world.

Results

During wheat straw fermentation, Lentinus edodes HAI 858 produced the most active Mn-dependent and Mn-independent peroxidases (1443.2 U L⁻¹ and 1045.5 U L⁻¹, respectively), while Pleurotus eryngii HAI 711 was the best laccase producer (7804.3 U L⁻¹). Visualized bends on zymogram confirmed these activities and demonstrated that laccases were the dominant ligninolytic enzymes in the studied species. Ganoderma lucidum BEOFB 435 showed considerable ability to degrade lignin (58.5%) and especially hemicellulose (74.8%), while the cellulose remained almost intact (0.7%). Remarkable selectivity in lignocellulose degradation was also noted in Pleurotus pulmonarius HAI 573 where degraded amounts of lignin, hemicellulose and cellulose were in ratio of 50.4%:15.3%:3.8%.

Conclusions

According to the presented results, it can be concluded that white-rot fungi, due to ligninolytic enzymes features and degradation potential, could be important participants in various biotechnological processes including biotransformation of lignocellulose residues/wastes in food, feed, paper and biofuels.

     

Transformation of14C labelled plant components in soil in relation to immobilization and remineralization of15N fertilizer

Summary Uniformly¹⁴C labelled glucose, cellulose and wheat straw and specifically¹⁴C labelled lignin component in corn stalks were aerobically incubated for 12 weeks in a chernozem soil alongwith¹⁵N labelled ammonium sulphate. Glucose was most readily decomposed, followed in order by cellulose, wheat straw and corn stalk lignins labelled at methoxyl-, side chain 2-and ring-C. More than 50% of¹⁴C applied as glucose, cellulose and wheat straw evolved as CO₂ during the first week. Lignin however, decomposed relatively slowly. A higher proportion of¹⁴C was transformed into microbial biomass whereas lignins contributed a little to this fraction.After 12 weeks of incubation nearly 60% of the lignin¹⁴C was found in humic compounds of which more than 70% was resistant to hydrolysis with 6N HCl. Maximum incorporation of¹⁵N in humic compounds was observed in cellulose amended soil. However, in this case more than 80% of the¹⁵N was in hydrolysable forms.Immobilization-remineralization of applied¹⁵N was most rapid in glucose treated soil and a complete immobilization followed by remineralization was observed after 3 days. The process was much slow in soil treated with cellulose, wheat straw or corn stalks. More than 70% of the newly immobilized N was in hydrolysable forms mainly reepresenting the microbial component.Serial hydrolysis of soil at different incubation intervals showed a greater proportion of 6N HCl hydrolysable¹⁴C and¹⁵N in fractions representing microbial material.¹⁴C from lignin carbons was relatively more uniformly distributed in different fractions as compared to glucose, cellulose and wheat straw where a major portion of¹⁴C was in easily hydrolysable fractions.     

Production of Manganese Peroxidase and Organic Acids and Mineralization of 14C-Labelled Lignin (14C-DHP) during Solid-State Fermentation of Wheat Straw with the White Rot Fungus Nematoloma frowardii

The basidiomycetous fungus Nematoloma frowardii produced manganese peroxidase (MnP) as the predominant ligninolytic enzyme during solid-state fermentation (SSF) of wheat straw. The purified enzyme had a molecular mass of 50 kDa and an isoelectric point of 3.2. In addition to MnP, low levels of laccase and lignin peroxidase were detected. Synthetic ¹⁴C-ring-labelled lignin (¹⁴C-DHP) was efficiently degraded during SSF. Approximately 75% of the initial radioactivity was released as ¹⁴CO₂, while only 6% was associated with the residual straw material, including the well-developed fungal biomass. On the basis of this finding we concluded that at least partial extracellular mineralization of lignin may have occurred. This conclusion was supported by the fact that we detected high levels of organic acids in the fermented straw (the maximum concentrations in the water phases of the straw cultures were 45 mM malate, 3.5 mM fumarate, and 10 mM oxalate), which rendered MnP effective and therefore made partial direct mineralization of lignin possible. Experiments performed in a cell-free system, which simulated the conditions in the straw cultures, revealed that MnP in fact converted part of the ¹⁴C-DHP to ¹⁴CO₂ (which accounted for up to 8% of the initial radioactivity added) and ¹⁴C-labelled water-soluble products (which accounted for 43% of the initial radioactivity) in the presence of natural levels of organic acids (30 mM malate, 5 mM fumarate).     

Mineralisation of 14C-labelled synthetic lignin and ligninolytic enzyme activities of litter-decomposing basidiomycetous fungi

Within a screening program, 27 soil litter-decomposing basidiomycetes were tested for ligninolytic enzyme activities using agar-media containing 2,2′-azinobis(3-ethylbenzthiazoline-6-sulphonate), a humic acid or Mn²⁺ ions as indicator substrates. Most active species were found within the family Strophariaceae (Agrocybe praecox, Stropharia coronilla, S. rugosoannulata) and used for mineralisation experiments with a ¹⁴C-ring-labelled synthetic lignin (¹⁴C-DHP). The fungi mineralised around 25% of the lignin to ¹⁴CO₂ within 12 weeks of incubation in a straw environment; about 20% of the lignin was converted to water-soluble fragments. Mn-peroxidase was found to be the predominant ligninolytic enzyme of all three fungi in liquid culture and its production was strongly enhanced in the presence of Mn²⁺ ions. The results of this study demonstrate that certain ubiquitous litter-decomposing basidiomycetes possess ligninolytic activities similar to the wood-decaying white-rot fungi, the most efficient lignin degraders in nature. Received: 20 April 2000 / Received revision: 12 July 2000 / Accepted: 16 July 2000     

Lignin solubilisation by Thermomonospora mesophila

Summary Thermomonospora mesophila degraded [¹⁴C]lignin-labelled wheat lignocellulose to yield high molecular weight water-soluble products and a small amount of ¹⁴CO₂. Solubilisation of [¹⁴C]lignin was found to be extracellular and inducible by growth on lignocellulose (straw) and hemicellulose (xylan), but was not correlated with xylanase or cellulase production.The acid-precipitable product of straw degradation by T. mesophila was found to be a complex of lignin, pentose-rich carbohydrate and protein with some similarity to humic acids. Solid-state ¹³C-NMR spectra of the dried product were generally similar to those of chemically extracted milled straw lignin but showed an increased content of carbonyl groups.The relationship between degradation and solubilisation of lignin is discussed and a role suggested for actinomycetes in humification and the exploitation of lignocellulose bioconversion.     

Effect of substrate nitrogen on lignin degradation by Pleurotus ostreatus

In order to determine the effect of substrate nitrogen (N) on ligninolytic activity, Pleurotus ostreatus was grown in solid media containing either growth-limiting (1mM) or excess (10mM) NH₄Cl. After 25 days, ¹⁴C–CO₂ production from ¹⁴C-cornstover lignin in low-N medium was 3 times that in nitrogen (N)-rich medium. Supplementation of low-N medium with glucose (0.3%) further enhanced ligninolytic activity. Decolorization of an aromatic, polymeric dye, Poly R-481, in solid media was also greatest under N-limiting conditions.     

Bioconversion of sugar cane crop residues with white-rot fungiPleurotus sp.

Four mushroom strains ofPleurotus spp. were cultivated on sugar cane crop residues for 30 days at 26°C. Biochemical changes affected the substrate as a result of fungal growth, in terms of nitrogen, lignin, cellulose and hemicellulose contents. All strains showed a strong ligninolytic activity together with variable cellulolytic and xylanolytic action.Pleurotus sajor-caju attacked lignin and cellulose at the same rate, showing a degradation of 47% and 55%, respectively. A better balance was shown by theP. ostreatus-P. pulmonarius hybrid, which exhibited the poorest cellulolytic action (39%) and the highest ligninolytic activity (67%). The average composition of mushroom fruit bodies, in terms of nitrogen, carbohydrates, fats and amino acid profiles, was determined. Crude protein and total carbohydrate varied from 23% to 33% and 36% to 68% of dry matter, respectively. Fat ranged from 3.3% to 4.7% and amino acid content from 12.2% to 22.2%. Slight evidence for a nitrogen fixing capability was encountered in the substrate to fruit body balance.     

Biodegradation of wheat straw lignocarbohydrate complexes (LCC)

Summary In laboratory and semi-industrial scale experiments the influence of the substrate water content, temperature, and incubation time on the progress of solid state fermentation of straw colonized by white rot fungi was investigated. The parameters used to evaluate the fermentation process were degradation of total organic matter and lignin, in vitro digestibility, the content of water soluble substances in the substrate and the pH.The degradation of total organic matter was species specific. Only Trametes hirsuta enhanced the degradation at elevated temperature (30 °C). With Abortiporus biennis, Ganoderma applanatum, and Pleurotus serotinus, elevated temperature had and adverse effect. Prolonged incubation only improved degradation of straw by the relatively slowgrowing fungi Ganoderma applanatum, Lenzites betulina, and Pleurotus sajor caju.Elevated temperature and prolonged incubation shifted the relative degradation rates in favour of total organic matter degradation. With Ganoderma applanatum, Pleurotus ostreatus, and Pleurotus serotinus lignin degradation, even on an absolute scale, was less at 30 °C than at 22 °C.In general, the in vitro digestibility also decreased, when the incubation time and temperature were raised. With Ganoderma applanatum the in vitro digestibility dropped below the value of the sterile straw control.Solid state fermentation of straw was at an optimum at a medium water content of 75 ml/25 g of substrate. However, most of the fungi tested could digest straw over a wide range of water content. At higher water contents (125–150 ml/25 g of substrate) an increased production of aerial mycelium was observed.In semi-industrial batch experiments (40 kg) with Abortiporus biennis the in vitro digestibility dropped below the reference value for sterile straw during the first 19 days of incubation. Later, the in vitro digestibility again rose and reached its optimum after about 60 days. The in vitro digestibility in the semi-industrial experiments was always lower than in the laboratory experiments (+9% and +25%, respectively).In long term experiments (2.5 kg batches, 8 months of incubation) very different values for the in vitro digestibility were found, and these depended on the fungus used (Abortiporus biennis, +16%; Pleurotus ostreatus, +4%; and Ganoderma applanatum, –27%).     

Substrate-dependent degradation patterns in the decay of wheat straw and beech wood by ligninolytic fungi

Summary The ability of 45 fungal strains to degrade wheat straw and beech wood was studied. Degradation patterns were defined in terms of chemical evolution of substrates and changes in lignin and polysaccharides. Trametes versicolor produced an important degradation of lignin and increased substrate digestibility, but it caused high weight losses and gave rise to similar decay patterns on both substrates. A preferential degradation of lignin was produced during straw transformation by Pleurotus eryngii. The increase of soluble lignin and decreases of lignin content and H/C ratio defined the degradation tendency after principal component analysis. The cation exchange capacity and water and alkali solubility presented the highest loading factors for the characterization of fungal transformation of beech wood. Offprint requests to: A. T. Martínez     

Influence of aromatic compounds on biodegradation of [14C]-labeled xylan and mannan by the white-rot fungus Phlebia radiata

Radiolabeled [¹⁴C]arabinoxylan from wheat meal and [¹⁴C]galactoglucomannan from red clover meal were prepared by using ¹⁴CO₂ as a precursor. Twice as much mannan was mineralized than xylan after 14 days of incubation with Phlebia radiata. Low-molecular-weight phenolic compounds structurally related to lignin increased during mineralization of both hemicellulose fractions. Veratryl alcohol increased degradation of arabinoxylan by approximately 28.5%, whereas veratric acid increased it by only 9.0%. Vanillic acid and ferulic acid also stimulated degradation by 16.6% and 34.7%, respectively. Veratryl alcohol and ferulic acid increased degradation of galactoglucomannan by approximately 75%. Veratraldehyde in both cases repressed the degradation process (23.6% arabinoxylan, 43.8% galactoglucomannan). These results indicate that the degradation of hemicelluloses, e.g., xylan and mannan, by P. radiata is enhanced by addition of aromatic compounds. Journal of Industrial Microbiology & Biotechnology (2002) 28, 168–172 DOI: 10.1038/sj/jim/7000221 Received 25 July 2001/ Accepted in revised form 23 October 2001     

Degradation of 14C-labelled poplar wood lignin by selected white-rot fungi

Summary Of eight white-rot fungi examined, seven fungi grew on nitrogen-limited poplar wood meal medium and degraded ¹⁴C-lignin in wood meal to ¹⁴CO₂. Increased oxygen enhanced both the rate and extent of degradation. However, whereas Pleurotus ostreatus, Pycnoporus cinnabarinus 115 and Pycnoporus cinnabarinus A-360 degraded 12–17% of ¹⁴C-(U)-lignin of poplar wood to ¹⁴CO₂ also in an air atmosphere, Sporotrichum pulverulentum, Phlebia radiata 79 and Phanerochaete sordida 37 degraded only 1–5% under these conditions. Addition of cellulose and glucose to the poplar wood medium stimulated degradation of ¹⁴C-(RING)-lignin of poplar wood by Phlebia radiata 79 but repressed degradation by Polyporus versicolor and Pleurotus ostreatus. Cellulose added to the wood meal medium had no effect on the degradation of lignin by Phanerochaete sordida 37 and Sporotrichum pulverulentum but glucose slightly repressed lignin degradation by these fungi. Those white-rot fungi which were considered as preferentially lignin attacking fungi could degrade ¹⁴C-(RING)-lignin of poplar wood efficiently under 100% oxygen. They did not require an extra energy source in addition to wood meal polysaccharides for rapid ring cleavage and they degraded up to 50–60% of the ¹⁴C-lignin to ¹⁴CO₂ in 6–7 weeks at a maximum rate of 3–4% per day.These results were reported in part at the Journées Internationales d'Etudes du Groupe Polyphenols, 29. 9.–1. 10. 1982, Université Paul Sabatier, Toulouse, France     

Simultaneous Pretreatment and Biohydrogen Production from Wheat Straw by Newly Isolated Ligninolytic <Emphasis Type="Italic">Bacillus</Emphasis> Sp. Strains with Two-Stage Batch Fermentation System

Biodegradation of agribiomass especially wheat straw to biohydrogen and biomethane is an encouraging approach to the current waste management problem. To do so, the biomass must first be pretreated to break down lignin thereby increasing accessibility of the substrate to fermentative organisms. In the current study, out of 20 isolates from the granular sludge of full-scale anaerobic digester, four ligninolytic Bacillus sp. strains were selected based on their lignin and Azure B degradation. Further, among the four isolates, Brevibacillus agri AN-3 exhibited the highest of 88.4 and 78.1% decrease in COD of lignin and Azure B respectively. These strains were also found to secrete optimum yields of lignin peroxidase (LiP) at pH 3, laccase (Lac) at pH 5, and xylanase and cellulase enzymes at pH 7. The strains demonstrated maximum activity of Lip and Lac at 50 °C and xylanase and cellulase at 60 °C after 72-h growth. Among the four strains, Brevibacillus agri AN-3 showed hydrogen (H₂) yield of 1.34 and 2.9 mol-H₂/mol from xylose and cellulose respectively. In two-phase wheat straw batch fermentation, Brevibacillus agri AN-3 produced 88.3 and 283.7 mL/gVS cumulative H₂ and methane (CH₄) respectively. Biotreatment with ligninolytic Bacillus sp. strains perceived that 261.4% more methane yield could be obtained from the wheat straw than using the untreated wheat straw in batch fermentation. This is the first study establishing not only the hydrogen potential of ligninolytic Bacillus sp. strains but also indicates a vital role of these species in developing standard inoculum and a biocatalyst for processing agribiomass.

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Graphical Abstract ?

     

The effect of white-rot basidiomycetes on chemical composition andin Vitro digestibility of wheat straw

Five white-rot basidiomycetes were evaluated for their potential to improve ruminal degradation of wheat straw.Polyorus brumalis, Lyophyllum ulmarium III,Trametes gibbosa, Pleurotus ostreatus, and aPleurotus ostreatus mutant were incubated on wheat straw for 30 d at 28°C. Detergent fiber, crude protein andin vitro dry matter digestibility (IVDMD) were determined. The results showed increasing crude protein and ash contents in fungus-treated straw. IVDMD values were increased in straws treated withP. ostreatus, P. ostreatus mutant andT. gibbosa only. Relative to untreated wheat straw the detergent fiber content—neutral detergent fiber (NDF), and acid detergent fiber (ADF) was reduced in fungus-treated straw and out of three fractions—hemicellulose, cellulose and lignin, hemicellulose showed the largest proportionate loss whereas lignin the smallest one in all 5 samples of fungus treated straw.     

13C NMR cross polarization and magic angle spinning (CPMAS) and gas chromatography/mass spectrometry analysis of the products from a soda pulp mill effluent decolourised by two Streptomyces strains

Two Streptomyces strains, UAH 30 and UAH 51, have been shown to decolourise a paper-mill effluent obtained after semichemical alkaline pulping of wheat straw. Fractionation of the effluent decolourised by strains UAH 30 and UAH 51 showed that 60% and 80% respectively of the alkali-lignin fraction have been removed from the effluent after 7 days of growth. ¹³C NMR cross polarization and magic angle spinning (CPMAS) spectra of the alkali-lignin remaining in the effluent after decolourisation revealed a decrease in the relative amount of aromatic lignin units compared to that obtained from the untreated effluent along with a reduction in the ratio of syringyl:guaiacyl units. Gas chromatography/mass spectrometry analysis of the low-molecular-mass compounds extracted from the decolourised effluent revealed the presence of new aromatic lignin-related compounds that were not present in the untreated control effluent. This was linked to a general depolymerization of larger lignin molecules occurring during decolourisation by the two Streptomyces strains. Identification of low-molecular-mass aromatic compounds extracted from the decolourised effluent revealed only the presence of p-hydroxyphenyl units in effluents decolourised by the strain UAH 30 while p-hydroxyphenyl, guaiacyl and syringyl units were detected in effluents decolourised by Streptomyces strain UAH 51. The study indicates that, while decolourisation is a common feature of the two Streptomyces strains, the mechanisms involved in the degradation of the lignin fractions may be different and strain-specific. Received: 8 July 1996 / Received revision: 9 October 1996 / Accepted: 14 October 1996     

Enhanced production of ligninolytic enzymes and decolorization of molasses distillery wastewater by fungi under solid state fermentation

Selected isolates of fungi were grown on wheat straw and corncob in the presence of different moistening agents such as water, molasses, potato dextrose broth and distillery effluent. All the fungal isolates responded differently with respect to growth and ligninolytic enzyme production. Fungal growth on different substrates was checked by calculating ergosterol content, which varied widely within a single species when grown on different substrates. The maximum laccase production was obtained for Aspergillus flavus TERI DB9 grown on wheat straw with molasses. For manganese peroxidase, highest production was in Aspergillus niger TERI DB20 grown on corncob with effluent. Among the two isolates positive for lignin peroxidase, the highest production was in Fusarium verticillioides ITCC 6140. This immobilized fungal biomass was then used for decolorization of effluent from a cane molasses based distillery. Maximum decolorization (86.33%) was achieved in Pleurotus ostreatus (Florida) Eger EM 1303 immobilized on corncob with molasses in a period of 28 days.     

Effect of three strains of Pleurotus tuber-regium (Fr.) Sing. on chemical composition and rumen fermentation of wheat straw

This study was conducted to investigate changes in in vitro dry matter digestibility (IVDMD) and cell wall constituent degradation in wheat straw treated with 3 strains of the fungus Pleurotus tuber-regium (PT). The incubation of wheat straw for 30 days at 28 degrees C improved IVDMD from 30.3% (UWS-untreated wheat straw) to 47.1% for strain PT1, to 48.5% for PT4, and was unchanged IVDMD-29.9% -for PT5. The growth of fungi was accompanied by the dry matter loss of wheat straw: 31.5% for PT1, 20.9% for PT4, and 4.8% for PT5. Fungal treatment was characterized by increased crude protein and ash contents (%) in all fungi-treated straws and reduced hemicellulose and lignin content. It is evident that enzymes of all 3 PT strains preferentially degraded hemicellulose and lignin over cellulose. Wheat straw treated with PT1 (TWS-PT1), PT4 (TWS-PT4), and PT5 (TWS-PT5) and barley (80% : 20%) were used as the experimental diets at the fermentation in the artificial rumen. UWS with barley (80% : 20%) served as the control diet. The fermentation of experimental diets was accompanied with increased IVDMD and a very low degree of hemicellulose degradation. Total gas and methane productions were similar in all diets. Moreover, total volatile fatty acid (VFA) production (mmol day(-1)), mol % of acetate, propionate, butyrate, isobutyrate, and isovalerate were not influenced during the fermentation of experimental diets. From the stoichiometric relations, production, utilization, and recovery of metabolic hydrogen and organic matter fermented were unchanged. Only the recovery of metabolic hydrogen in TWS-PT5 was significantly increased in comparison to control diet. Total microbial production showed the tendency of lower values in experimental diets, and it was accompanied with a significant decrease of ammonia nitrogen (mg L(-1)). Finally the results showed that the strains of Pleurotus tuber-regium can improve the quality of wheat straw, but the loss of dry matter (DM) (mainly hemicellulose) limits the effective utilization of fungi-treated straw in ruminant digestion.     

Ligninolytic system of Phanerochaete chrysosporium: Inhibition by o-Phthalate

The degradation rate of [synthetic-¹⁴C]-lignin to ¹⁴CO₂ by Phanerochaete chrysosporium in cultures buffered with 0.01 M 2,2-dimethylsuccinate (DMS) was twice that in 0.01 M o-phthalate-buffered cultures. This difference could be totally accounted for by o-phthalate inhibition of the activity of the ligninolytic system. ¹⁴CO₂ production from ring-, sidechain-, and methoxyl-labeled lignins was inhibited, the degree of inhibition being dependent on o-phthalate concentration. Oxidations of ¹⁴C-glucose, ¹⁴C-acetovanillone, and ¹⁴C-apocynol were not inhibited; thus o-phthalate is not a general inhibitor, and might inhibit activities involved in attack of the lignin polymer. DMS is a suitable buffer for the ligninolytic system. Degradation rates of ring-labeled lignin to ¹⁴CO₂ of 10–15% in 24 h were obtained consistently over the pH range 3.6–4.5, with an optimum near pH 4.0.Non-Standard Abbreviations DMS dimethylsuccinate     

Thermophilic Anaerobic Biodegradation of [14C]Lignin, [14C]Cellulose, and [14C]Lignocellulose Preparations

Thermophilic (55°C) anaerobic enrichment cultures were incubated with [¹⁴C-lignin]lignocellulose, [¹⁴C-polysaccharide]lignocellulose, and kraft [¹⁴C]lignin prepared from slash pine, Pinus elliottii, and ¹⁴C-labeled preparations of synthetic lignin and purified cellulose. Significant but low percentages (2 to 4%) of synthetic and natural pine lignin were recovered as labeled methane and carbon dioxide during 60-day incubations, whereas much greater percentages (13 to 23%) of kraft lignin were recovered as gaseous end products. Percentages of label recovered from lignin-labeled substrates as dissolved degradation products were approximately equal to percentages recovered as gaseous end products. High-pressure liquid chromatographic analyses of CuO oxidation products of sound and degraded pine lignin indicated that no substantial chemical modifications of the remaining lignin polymer, such as demethoxylation and dearomatization, occurred during biodegradation. The polysaccharide components of pine lignocellulose and purified cellulose were relatively rapidly mineralized to methane and carbon dioxide; 31 to 37% of the pine polysaccharides and 56 to 63% of the purified cellulose were recovered as labeled gaseous end products. An additional 10 to 20% of the polysaccharide substrates was recovered as dissolved degradation products. Overall, these results indicate that elevated temperatures can greatly enhance rates of anaerobic degradation of lignin and lignified substrates to methane and low-molecular-weight aromatic compounds.     

Manganese-Mediated Lignin Degradation by Pleurotus pulmonarius

Pleurotus pulmonarius produced the strongest degradation of lignin during solid-state fermentation of [(sup14)C]lignin wheat straw with different fungi. A manganese-oxidizing peroxidase seemed to be involved in lignin attack, since the addition of Mn(sup2+) to the culture increased lignin mineralization by ca. 125%. This enzyme was purified and characterized from both solid-state fermentation and liquid cultures.