Depolymerization and decolorization of kraft lignin by bacterium Comamonas sp. B-9

There is no commercial or industrial-scale process for the remediation of black liquor using microorganisms to date. One of the most important causes is that most microorganisms are not able to use lignin as their principal metabolic carbon or energy source. The bacterial strain Comamonas sp. B-9 has shown remarkable ability to degrade kraft lignin and decolorize black liquor using lignin as its principal metabolic carbon and energy source. This report looks at the depolymerization and decolorization of kraft lignin by Comamonas sp. B-9. The degradation, decolorization, and total carbon removal reached 45, 54, and 47.3 %, respectively, after 7 days treatment. Comamonas sp. B-9 was capable of depolymerizing kraft lignin effectively as analyzed by gel permeation chromatography and decolorization via degrading benzene ring structures as shown using Fourier transform infrared spectroscopy analysis.     

Biodegradation of kraft-lignin by <Emphasis Type="Italic">Bacillus</Emphasis> sp. isolated from sludge of pulp and paper mill

Eight bacterial strains were isolated on kraft lignin (KL) containing mineral salt medium (L-MSM) agar with glucose and peptone from the sludge of pulp and paper mill. Out of these, ITRC-S8 was selected for KL degradation, because of its fast growth at highest tested KL concentration and use of various lignin-related low molecular weight aromatic compounds (LMWACs) as sole source of carbon and energy. The bacterium was identified by biochemical tests as Gram-positive, rod-shaped and non-motile. Subsequent 16S rRNA gene sequencing showed 95% base sequence homology and it was identified as Bacillus sp. In batch experiments, a decrease in pH was observed initially followed by an increase till it reached an alkaline pH, which did not alter the culture growth significantly. The bacterium reduced the colour and KL content of 500 mg l⁻¹ KL in MSM, in the presence of glucose and peptone, at pH 7.6, temperature 30°C, agitation of 120 rpm and 6 days of incubation by 65 and 37% respectively. Significant reduction in colour and KL content in subsequent incubations is indicative of a co-metabolism mechanism, possibly due to initial utilization of added co-substrates for energy followed by utilization of KL as a co-metabolic. The degradation of KL by bacterium was confirmed by GC-MS analysis indicating formation of several LMWACs such as t-cinnamic acid, 3, 4, 5-trimethoxy benzaldehyde and ferulic acid as degradation products, which were not present in the control (uninoculated) sample. This favours the idea of biochemical modification of the KL polymer to a single monomer unit.     

Biodegradation of kraft lignin by a newly isolated bacterial strain, <Emphasis Type="Italic">Aneurinibacillus aneurinilyticus</Emphasis> from the sludge of a pulp paper mill

A kraft lignin-degrading bacterium (ITRC S ₇ ) was isolated from sludge of pulp and paper mill and characterized as Aneurinibacillus aneurinilyticus by biochemical tests and 16SrRNA gene sequencing. The bacterium did not utilize kraft lignin (KL) as the sole source of carbon and energy. However, this strain reduced the color (58%) and lignin content (43%) from kraft lignin-mineral salt medium when supplemented with glucose at pH 7.6 and 30°C after 6 days. The degradation on addition of glucose in culture medium is clear evidence of co-metabolism of KL by A. aneurinilyticus. The analysis of lignin degradation products by GC-MS in ethyl acetate extract from an A. aneurinilyticus-inoculated sample revealed the formation of low molecular weight aromatic compounds such as guaiacol, acetoguaiacone, gallic acid and ferulic acid, indicating that the bacterium can oxidize of the sinapylic (G units) and coniferylic (S units) alcohol units which are the basic moieties that build the hardwood lignin structure. The low molecular weight aromatic compounds identified in extracts of the inoculated sample favors the idea of biochemical modification of the KL to a single aromatic unit.     

Identification of low molecular weight aromatic compounds by gas chromatography–mass spectrometry (GC–MS) from kraft lignin degradation by three Bacillus sp.

Three bacterial strains identified as Paenibacillus sp., Aneurinibacillus aneurinilyticus and Bacillus sp. have been shown to decolourise kraft lignin in 6 days of incubation. The release of low molecular weight aromatic compounds by these bacterial strains during degradation of kraft lignin was analysed by GC–MS analysis. The total ion chromatograph (TIC) of ethyl acetate extract from kraft lignin sample inoculated by Paenibacillus sp. was similar to control except some minor changes in the chromatographic profile indicating incapability of this bacterium to modify kraft lignin. On the other hand the TIC of ethyl acetate extract from kraft lignin inoculated by A. aneurinilyticus and Bacillus sp. caused formation of several aromatic lignin-related compound that were not present in the extract of control. The compounds identified in extract of the sample degraded by A. aneurinilyticus were guaiacol, acetoguiacone, gallic acid and ferulic acid while t-cinnamic acid, 3,4,5 trimethoxy benzaldehyde, and ferulic acid by Bacillus sp. indicating oxidization of coniferylic (G units) and sinapylic (S units) alcohol of lignin polymer. Differences between the identified compounds from different bacterial treatment were strain-specific. Among the identified aromatic compounds, ferulic acid and 3,4,5-trimethoxy benzaldehyde could be useful to the industry of preservatives, aromas and perfumes.     

废次烟叶提取液源木质素降解菌Bacillus subtilis SM降解特性

【目的】目前造纸法再造烟叶工艺已经成为我国重要的废烟叶处理和利用方式,该工艺中烟梗中高木质素的降解是个挑战性的需解决问题。从废次烟叶提取液(Tobacco waste extract,TWE)中筛选木质素的降解微生物用来直接处理烟梗或烟末提取液,可实现对木质素含量的调控。【方法】将废次烟叶提取液(TWE)浓缩液中分离出的Bacillus subtilis SM接种到以Kraft木质素为唯一碳源的无机盐培养基中,在pH 7.0、30°C培养基中培养4 d来检测菌株对木质素的降解效果。通过HPLC、TOC、GPC和色度来表征SM对木质素的降解,并采用烟梗无机盐培养基在pH 7.0、30°C培养4 d检测SM对烟梗木质素的降解。【结果】HPLC结果显示SM在以木质素磺酸钠为唯一碳源的无机盐培养基中可全部降解分子质量为534.5的木质素磺酸钠,而对Kraft木质素降解不明显,仅观察到组分的变化。脱色结果显示脱色率达到40.7%,但在对Kraft木质素矿化方面矿化率只能达到5.4%。SM在烟梗无机盐培养基中可使烟梗失重率分别达到50%以上(对照组为18.9%),烟梗中木质素含量减少了70%左右。【结论】来源于废次烟叶提取液(TWE)的Bacillus subtilis SM能够以Kraft木质素为唯一碳源生长,也能够有效降解烟梗中的木质素,可应用于烟草废弃物原料中木质素的降解。     

Effect ofStreptomyces viridosporus T7A on kraft lignin

Summary The ability of the lignino-cellulolytic actinomyceteStreptomyces viridosporus T7A to attack purified fractions of kraft lignin was examined. In the presence of 0.3% yeast extract, high-molecular weight kraft lignin (MW>3000, ether-insoluble fraction) does not affect growth of this microorganism significantly, whereas low-molecular weight kraft lignin (MW<3000, ether-soluble fraction) inhibits its development. Accordingly, average molecular weight of the ether-insoluble fraction after bacterial growth remained unaltered, as measured by Sephadex G-50 gel permeation chromatography. Slight modifications were detected by high performance liquid chromatography in the ether-soluble fraction after incubation with the microorganism.S. viridosporus T7A partially decolorized Remazol Brilliant Blue R during growth on wheat lignocellulose. However, decolorization of either fraction of kraft lignin was not observed. These results suggest that the filamentous bacteriumS. viridosporus T7A is not suitable for pulp mill effluent treatment.     

Isolation and characterization of lignin‐degrading bacteria from rainforest soils

The deconstruction of lignin to enhance the release of fermentable sugars from plant cell walls presents a challenge for biofuels production from lignocellulosic biomass. The discovery of novel lignin‐degrading enzymes from bacteria could provide advantages over fungal enzymes in terms of their production and relative ease of protein engineering. In this study, 140 bacterial strains isolated from soils of a biodiversity‐rich rainforest in Peru were screened based on their oxidative activity on ABTS, a laccase substrate. Strain C6 (Bacillus pumilus) and strain B7 (Bacillus atrophaeus) were selected for their high laccase activity and identified by 16S rDNA analysis. Strains B7 and C6 degraded fragments of Kraft lignin and the lignin model dimer guaiacylglycerol‐β‐guaiacyl ether, the most abundant linkage in lignin. Finally, LC–MS analysis of incubations of strains B7 and C6 with poplar biomass in rich and minimal media revealed that a higher number of compounds were released in the minimal medium than in the rich one. These findings provide important evidence that bacterial enzymes can degrade and/or modify lignin and contribute to the release of fermentable sugars from lignocellulose. Biotechnol. Bioeng. 2013; 110: 1616–1626. © 2013 Wiley Periodicals, Inc.     

Biodegradation of nicotine by a newly isolated Agrobacterium sp. strain S33

Aims: To isolate and characterize bacteria capable of degrading nicotine from the rhizospheric soil of a tobacco plant and to use them to degrade the nicotine in tobacco solid waste. Methods and Results: A bacterium, strain S33, was newly isolated from the rhizospheric soil of a tobacco plant, and identified as Agrobacterium sp. based on morphology, physiological tests, Biolog MicroLog3 4·20 system and 16S rRNA gene sequence. Using nicotine as the sole source of carbon and nitrogen in the medium, it grew optimally with 1·0 g l^?1 of nicotine at 30°C and pH 7·0, and nicotine was completely degraded within 6 h. The resting cells prepared from the glucose‐ammonium medium or LB medium could not degrade nicotine within 10 h, while those prepared from the nicotine medium could completely degrade 3 g l^?1 of nicotine in 1·5 h at a maximal rate of 1·23 g nicotine h^?1 g^?1 dry cell. Using the medium containing nicotine, glucose and ammonium simultaneously to cultivate strain S33, the resting cells could degrade 98·87% of nicotine in tobacco solid waste with the concentration as 30 mg nicotine g^?1 dry weight tobacco solid waste within 7 h at a maximal rate of 0·46 g nicotine h^?1 g^?1 dry cell. Conclusions: This is the first report that Agrobacterium sp. has the ability to degrade nicotine. Agrobacterium sp. S33 could use nicotine as the sole source of carbon and nitrogen. The use of resting cells of the strain S33 prepared from the nicotine–glucose–ammonium medium was an effective method to degrade nicotine and detoxify tobacco solid waste. Significance and Impact of the Study: Nicotine in tobacco wastes is both toxic and harmful to human health and the environment. This study showed that Agrobacterium sp. S33 may be suitable for the disposal of tobacco wastes and reducing the nicotine content in tobacco leaves.     

Isolation and characterization of new poly(3HB)‐accumulating star‐shaped cell‐aggregates‐forming thermophilic bacteria

Aims: This study aimed at isolating thermophilic bacteria that utilize cheap carbon substrates for the economically feasible production of poly(3‐hydroxybutyrate), poly(3HB), at elevated temperatures. Methods and Results: Thermophilic bacteria were enriched from an aerobic organic waste treatment plant in Germany, and from hot springs in Egypt. Using the viable colony staining method for hydrophobic cellular inclusions with Nile red in mineral salts medium (MSM) containing different carbon sources, six Gram‐negative bacteria were isolated. Under the cultivation conditions used in this study, strains MW9, MW11, MW12, MW13 and MW14 formed stable star‐shaped cell‐aggregates (SSCAs) during growth; only strain MW10 consisted of free‐living rod‐shaped cells. The phylogenetic relationships of the strains as derived from 16S rRNA gene sequence comparisons revealed them as members of the Alphaproteobacteria. The 16S rRNA gene sequences of the isolates were very similar (>99% similarity) and exhibited similarities ranging from 93 to 99% with the most closely related species that were Chelatococcus daeguensis, Chelatococcus sambhunathii , Chelatococcus asaccharovorans, Bosea minatitlanensis, Bosea thiooxidans and Methylobacterium lusitanum. Strains MW9, MW10, MW13 and MW14 grew optimally in MSM with glucose, whereas strains MW11 and MW12 preferred glycerol as sole carbon source for growth and poly(3HB) accumulation. The highest cell density and highest poly(3HB) content attained were 4·8 g l^?l (cell dry weight) and 73% (w/w), respectively. Cells of all strains grew at temperatures between 37 and 55°C with the optimum growth at 50°C. Conclusions: New PHA‐accumulating thermophilic bacterial strains were isolated and characterized to produce poly(3HB) from glucose or glycerol in MSM at 50°C. SSCAs formation was reported during growth. Significance and Impact of the Study: To the best of our knowledge, this is the first report on the formation of SSCAs by PHA‐accumulating bacteria and also by thermophilic bacteria. PHA‐producing thermophiles can significantly reduce the costs of fermentative PHA production.     

Enhanced degradation of lignin by the strain fusion of Pseudomonas putida and Gordonia sp

The protoplast fusion technique was applied to construct a more efficient engineering microbial strain to degrade lignin by fusing two strains, Pseudomonas putida and Gordonia sp. At an initial lignin concentration of 900?mg/L, COD, BOD, TOC removal efficiencies increased from 69–76%, 69–72%, and 70–72% by the parent stains to 83%, 83%, and 83% of the fused strain, respectively. IR and HPLC analyses of the treated solution suggested that the fused strains were more capable of breaking the C_α–C_β bonds of the benzene ring in lignin compared to its parent strains, yielding syringyls as the main product. GC–MS analysis was used to identify the release of three-types of lower molecular intermediates: ring-opening, monomer, and dipolymer products. The phenolic hydroxyl group in lignin was oxidized to carbonyls, followed by further degradation to acids and esters. The carboxyl group on the ether linkage that maintains the macromolecular structure of lignin was oxidized to acyls, which further led to depolymerization and the opening of benzene ring.     

Recovery of kraft lignin from pulping wastewater via emulsion liquid membrane process

Kraft lignin (KL) is a renewable source of many valuable intermediate biochemical products currently derived from petroleum. An excessive of lignin comes from pulping wastewater caused an adverse pollution problems hence affecting human and aquatic life. A comprehensive study pertaining to emulsion liquid membrane (ELM) extraction of lignin from pulping wastewater was presented. ELM formulation contains Aliquat 336 as carrier, kerosene as diluent, sodium bicarbonate (NaHCO₃) as stripping agent and Span 80 as surfactant. The emulsion stability was investigated at different surfactant concentrations, homogenizer speed and emulsification time. Modifier (2‐ethyl‐1‐hexanol) was added to avoid segregation of third phase while improving the emulsion stability. At optimum conditions, 95% and 56% of lignin were extracted and recovered, respectively at 10 min of extraction time, 0.007 M of Aliquat 336, 0.1 M of NaHCO₃ and 1:5 of treat ratio. Additional of modifier was contributed to highest recovery up to 98%. The ELM process was found to be equally feasible and quite effective in the recovery of KL from real pulping wastewater. Therefore, ELM process provides a promising alternative technology to recover KL from pulping wastewater while solving the environmental problems simultaneously. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1305–1314, 2015     

The ability of white-rot fungi to degrade the endocrine-disrupting compound nonylphenol

Phanerochaete chrysosporium, Pleurotus ostreatus, Trametes versicolor and Bjerkandera sp. BOL13 were tested for their ability to degrade the endocrine-disrupting compound nonylphenol at an initial concentration of 100 mg l^–1. The highest removals were achieved with T. versicolor and Bjerkandera sp. BOL13, which were able to degrade 97 mg l^–1 and 99 mg l^–1 of nonylphenol in 25 days of incubation, respectively. Nonylphenol removal was associated with the production of laccase by T. versicolor, but the levels of laccase, manganese peroxidase and lignin peroxidase produced by Bjerkandera sp. BOL13 were very low. At 14°C, T. versicolor and Bjerkandera sp. BOL13 sustained the removal of 88 mg l^–1 and 79 mg l^–1 of nonylphenol, respectively. No pollutant removal was recorded at 4°C, although both fungi could grow at this temperature in the absence of nonylphenol. A microtoxicity assay showed that the fungi produced compounds that were toxic to Vibrio fischerii; and thus a reduction in toxicity could not be correlated with nonylphenol metabolism. T. versicolor and Bjerkandera sp. BOL13 were capable of colonizing soil artificially contaminated with 430 mg kg^–1 of nonylphenol. Only 1.3±0.1% of nonylphenol remained in the soil after 5 weeks of incubation.     

Molybdate reduction by Pseudomonas sp. strain DRY2

Aims: To isolate and characterize a potent molybdenum‐reducing bacterium. Methods and Results: A minimal salt medium supplemented with 10 mmol l^?1 molybdate, glucose (1·0%, w/v) as a carbon source and ammonium sulfate (0·3%, w/v) as a nitrogen source was used in the screening process. A molybdenum‐reducing bacterium was isolated and tentatively identified as Pseudomonas sp. strain DRY2 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny. Strain DRY2 produced 2·4, 3·2 and 6·2 times more molybdenum blue compared to Serratia marcescens strain DRY6, Enterobacter cloacae strain 48 and Eschericia coli K12, respectively. Molybdate reduction was optimum at 5 mmol l^?1 phosphate. The optimum molybdate concentration that supported molybdate reduction at 5 mmol l^?1 phosphate was between 15 and 25 mmol l^?1. Molybdate reduction was optimum at 40°C and at pH 6·0. Phosphate concentrations higher than 5 mmol l^?1 strongly inhibited molybdate reduction. Inhibitors of electron transport system such as antimycin A, rotenone, sodium azide and cyanide did not inhibit the molybdenum‐reducing enzyme activity. Chromium, copper, mercury and lead inhibited the molybdenum‐reducing activity. Conclusions: A novel molybdenum‐reducing bacterium with high molybdenum reduction capacity has been isolated. Significance and Impact of the Study: Molybdenum is an emerging global pollutant that is very toxic to ruminants. The characteristics of this bacterium suggest that it would be useful in the bioremediation of molybdenum pollutant.     

Bacterial degradation of dehydropolymers of coniferyl alcohol

A bacterial isolate identified as Xanthomonas sp. proved to be ligninolytic due to its ability to degrade ¹⁴C-labeled dehydropolymers of coniferyl alcohol (DHP) and [14C]lignocellulose complexes from corn plants (Zea mays). Several parameters of ligninolysis were evaluated and it was shown that resting cells degrade DHP as sole carbon source. Enhancement of DHP degradation in the presence of ferulic acid or water-soluble fractions of DHP or of dioxane lignin from wheat was demonstrated. It is shown that a dissociation of DHP takes place during incubation in the absence of the bacteria which is reflected in a shift of DHP to lower molecular weight fractions. Bacterial degradation of [14C] DHP results in the release of ¹⁴CO₂ and in the incorporation of the ¹⁴C-label into the biomass of the bacteria, as shown by chemical and biological methods.Abbreviations Bq Becquerel, measure for radioactivity according to SI nomenclature - DHP dehydropolymers of coniferyl alcohol - DMF dimethylformamide - DMSO dimethyl sulfoxide - HPLC high performance liquid chromatography - TCA trichloroacetic acid - THF tetrahydrofuran     

Biodegradation of naphthalene-2-sulfonic acid present in tannery wastewater by bacterial isolates <Emphasis Type="Italic">Arthrobacter</Emphasis> sp. 2AC and <Emphasis Type="Italic">Comamonas</Emphasis> sp. 4BC

Two bacterial strains, 2AC and 4BC, both capable of utilizing naphthalene-2-sulfonic acid (2-NSA) as a sole source of carbon, were isolated from activated sludges previously exposed to tannery wastewater. Enrichments were carried out in mineral salt medium (MSM) with 2-NSA as the sole carbon source. 16S rDNA sequencing analysis indicated that 2AC is an Arthrobacter sp. and 4BC is a Comamonas sp. Within 33 h, both isolates degraded 100% of 2-NSA in MSM and also 2-NSA in non-sterile tannery wastewater. The yield coefficient was 0.33 g biomass dry weight per gram of 2-NSA. A conceptual model, which describes the aerobic transformation of organic matter, was used for interpreting the biodegradation kinetics of 2-NSA. The half-lives for 2-NSA, at initial concentrations of 100 and 500 mg/l in MSM, ranged from 20 h (2AC) to 26 h (4BC) with lag-phases of 8 h (2AC) and 12 h (4BC). The carbon balance indicates that 75–90% of the initial TOC (total organic carbon) was mineralized, 5–20% remained as DOC (dissolved organic carbon) and 3–10% was biomass carbon. The principal metabolite of 2-NSA biodegradation (in both MSM and tannery wastewater) produced by Comamonas sp. 4BC had a MW of 174 and accounted for the residual DOC (7.0–19.0% of the initial TOC and 66% of the remaining TOC). Three to ten percent of the initial TOC (33% of the remaining TOC) was associated with biomass. The metabolite was not detected when Arthrobacter sp. 2AC was used, and a lower residual DOC and biomass carbon were recorded. This suggests that the two strains may use different catabolic pathways for 2-NSA degradation. The rapid biodegradation of 2-NSA (100 mg/l) added to non-sterile tannery wastewater (total 2-NSA, 105 mg/l) when inoculated with eitherArthrobacter 2AC or Comamonas 4BC showed that both strains were able to compete with the indigenous microorganisms and degrade 2-NSA even in the presence of alternate carbon sources (DOC in tannery wastewater = 91 mg/l). The results provide information useful for the rational design of bioreactors for tannery wastewater treatment.     

Selective Degradation of Wood Components by White-Rot Fungi

In order to find naturally occurring white-rot fungi which preferentially degrade lignin. 25 different species of such fungi were cultivated on pine wood blocks and on kraft lignin agar plates with and without cellulose. Due to differences in phenol oxidase reactions on the kraft lignin agar plates, the 25 fungi could be divided into two groups, 1 and 2, which also differed in other properties. The three Group I fungi Sporotrichum pulverulentum, Phanerochaete sp. L1 and Polyporus dichrous produced high levels of endo-l,4-β-glucanase and cellobiose:quinone oxidoreductase in shaking cellulose flasks and a low level of phenol oxidase in standing wood meal flasks, The four fungi Merulius tremellosus, Phlebia radiata, Pycuoporus cinnabarinus and Pleurotus ostreatus from Group 2, on the other hand, produced low levels of endo-1,4-β-glucanase and cellobiose:.quinone oxidoreductase in the cellulose. flasks and a high level of phenol oxidase in the wood meal flasks. Analyses of pine wood blocks degraded by the above-mentioned fungi in the presence of either malt extract, asparagine or NH₄H₂PO₄ revealed that malt extract gave good lignin degradation. In the presence of this nutrient source. P. cinnabarinus, at 3.4% weight loss, even degraded 12.5% lignin without loss of cellulose or mannan. No common degradation pattern was, however, obtained using mall extract, asparagine or NH₄H₂PO₄, It is suggested that while-rot fungi, which preferentially degrade lignin, may be found among Group 2 fungi producing large amounts of phenol oxidases.     

Biochemical investigation of kraft lignin degradation by Pandoraea sp. B-6 isolated from bamboo slips

Kraft lignin (KL) is the major pollutant in black liquor. The bacterial strain Pandoraea sp. B-6 was able to degrade KL without any co-substrate under high alkaline conditions. At least 38.2 % of chemical oxygen demand and 41.6 % of color were removed in 7 days at concentrations from 1 to 6 g L^?1. The optimum pH for KL degradation was 10 and the optimum temperature was 30 °C. The greatest activities of 2,249.2 U L^?1 for manganese peroxidase and 1,120.6 U L^?1 for laccase were detected on the third and fifth day at pH 10, respectively. Many small molecules, such as cinnamic acid, ferulic acid, 2-hydroxy benzyl alcohol, and vanillyl methyl ketone, were formed during the period of KL degradation based on GC–MS analysis. These results indicate that this strain has great potential for biotreatment of black liquor.     

The importance of phenol oxidase activity in lignin degradation by the white-rot fungus Sporotrichum pulverulentum

The lignin degradation abilities of wildtype, a phenol oxidase-less mutant and a phenol oxidase-positive revertant of Sporotrichum pulverulentum were compared to determine if phenol oxidase activity is necessary for lignin degradation by white-rot fungi. The phenol oxidase-less mutant was unable to degrade kraft lignin or wood. The phenol oxidase-positive revertant, however, regained the ability of the wildtype to degrade kraft lignin and all of the major components of wood. It was found that kraft lignin and lignin-related phenols decreased cellulase and xylanase production by the phenol oxidase-less mutant. Addition of highly purified laccase increased the production of endo-1,4--glucanase in the phenol oxidase-less mutant in the presence of vanillic acid and kraft lignin. After addition of laccase to kraft lignin agar plates, the phenol oxidase-less mutant could degrade kraft lignin.It is proposed that phenol oxidase function in regulating the production of both lignin-and polysaccharide-degrading enzymes by oxidation of lignin and lignin-related phenols when S. pulverulentum is growing on wood.Abbreviation WT wildtype Sporotrichum pulverulentum Research supported by a grant from Stiftelsen Nils and Dorthi Troëdssons forskningsfond     

Early attack and subsequent changes produced in an industrial lignin by a fungal laccase and a laccase-mediator system: an analytical approach

An industrial kraft pine lignin (Indulin AT, KL) was characterized and treated in both aqueous-buffered media and dioxane to water, either with a partially purified laccase from Fusarium proliferatum or with the laccase plus 2,2′-azino-bis-3-ethylbenzothiazoline-6-sulfonic-acid (ABTS) as mediator. The changes in the lignin after different incubation periods were analyzed through the application of high performance liquid chromatography (HPLC), UV–visible (Vis) spectroscopy and pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS). At the onset of incubation, laccase-treated samples showed a slight polymerization and strong modifications in UV–Vis spectra. Through Py-GC/MS, a decrease in phenolic and methoxy-bearing pyrolysis products was observed, in contrast to an increase in the more oxidized products. After longer incubation periods (48 h) a substantial polymerization was detected by HPLC, along with a decrease in the guaiacyl (G) units. In contrast, the analysis by HPLC of the samples recovered from the laccase-ABTS system (LMS) showed an intense depolymerization, accompanied by a sizeable loss in G units and a decrease in the methyl and ethyl side-chain phenolic compounds. These results provide conclusive evidence of a rapid initial attack of the industrial lignin by laccase and notable modifications in the KL after longer incubation periods with laccase or LMS.