Thiourea effect on the lignin biodegradation

Summary Lignin degradation by peroxidase or an extracellular crude ligninase in the presence of thiourea is enhanced. Veratryl alcohol, exhibits the same behaviour in both cases. Thiourea is proposed as an additive in biopulping     

Recent developments in the understanding of lignin biodegradation

This paper briefly describes the historical background, the biological nature, epidemiology, diagnosis, and the control of leptospira and leptospirosis (Weil's disease).     

Influence of lignin peroxidase concentration and localisation in lignin biodegradation by Phanerochaete chrysosporium

Summary The lignin mineralization rate in cultures of Phanerochaete chrysosporium increases with lignin peroxidase concentration up to 20 nkat ml^–1. At higher concentrations the rate of lignin mineralization decreases with increasing lignin peroxidase concentration. The amount of mycelium is not a limiting factor for lignin mineralization at high exocellular lignin peroxidase in association with the mycelium as pellets and no free exocellular enzyme induce a lignin mineralization rate equivalent to cultures reconstituted with washed pellets supplemented with 15 nkat ml^–1 of exogenous free enzyme. These results show that although lignin degradation by lignin peroxidase seems to be facilitated when lignin peroxidase is localised on the surface of the mycelium, free exocellular lignin peroxidase can also efficiently enhance mineralization of lignin by P. chrysosporium.     

Fungal biodegradation and enzymatic modification of lignin

Lignin, the most abundant aromatic biopolymer on Earth, is extremely recalcitrant to degradation. By linking to both hemicellulose and cellulose, it creates a barrier to any solutions or enzymes and prevents the penetration of lignocellulolytic enzymes into the interior lignocellulosic structure. Some basidiomycetes white-rot fungi are able to degrade lignin efficiently using a combination of extracellular ligninolytic enzymes, organic acids, mediators and accessory enzymes. This review describes ligninolytic enzyme families produced by these fungi that are involved in wood decay processes, their molecular structures, biochemical properties and the mechanisms of action which render them attractive candidates in biotechnological applications. These enzymes include phenol oxidase (laccase) and heme peroxidases [lignin peroxidase (LiP), manganese peroxidase (MnP) and versatile peroxidase (VP)]. Accessory enzymes such as H₂O₂-generating oxidases and degradation mechanisms of plant cell-wall components in a non-enzymatic manner by production of free hydroxyl radicals (·OH) are also discussed.     

Oxidation of monomethoxylated aromatic compounds by lignin peroxidase: role of veratryl alcohol in lignin biodegradation

Lignin peroxidase (LiP), an extracellular heme enzyme from the lignin-degrading fungus Phanerochaete chrysosporium, catalyzes the H2O2-dependent oxidation of a variety of nonphenolic lignin model compounds. The oxidation of monomethoxylated lignin model compounds, such as anisyl alcohol (AA), and the role of veratryl alcohol (VA) in LiP reactions were studied. AA oxidation reached a maximum at relatively low H2O2 concentrations, beyond which the extent of the reactions decreased. The presence of VA did not affect AA oxidation at low molar ratios of H2O2 to enzyme; however, at ratios above 100, the presence of VA abolished the decrease in AA oxidation. Addition of stoichiometric amounts of AA to LiP compound II (LiPII) resulted in its reduction to the native enzyme at rates that were significantly faster than the spontaneous rate of reduction, indicating that AA and other monomethoxylated aromatics are directly oxidized by LiP, albeit slowly. Under steady-state conditions in the presence of excess H2O2 and VA, a visible spectrum for LiPII was obtained. In contrast, under steady-state conditions in the presence of AA a visible spectrum was obtained for LiPIII*, a noncovalent complex of LiPIII and H2O2. AA competitively inhibited the oxidation of VA by LiP; the Ki for AA inhibition was 32 microM. Addition of VA to LiPIII* resulted in its conversion to the native enzyme. In contrast, AA did not convert LiPIII* to the native enzyme; instead, LiPIII* was bleached in the presence of AA. Thus, AA does not protect LiP from inactivation by H2O2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anaerobic biodegradation of the lignin and polysaccharide components of lignocellulose and synthetic lignin by sediment microflora

[This corrects the article on p. 998 in vol. 47.].     

Manganese, Mn-dependent peroxidases, and the biodegradation of lignin

Manganese and Mn-dependent peroxidases have been implicated in the enzymatic degradation of lignin. However, the specific role of manganese is uncertain. We report here the novel observation that in the absence of enzyme, suitably chelated Mn3+ is a ligninolytic agent capable of oxidizing veratryl alcohol, lignin model compounds, and lignin. We also demonstrate the unexpected effect of reducing agents which stimulate the oxidations by Mn3+. The stimulation is apparently through the production of a reduced oxygen species likely to be superoxide. These observations provide a fresh insight into the process of lignin biodegradation.     

Effect of steam explosion on biodegradation of lignin in wheat straw

The effect of steam explosion pretreatment on biodegradation of lignin in wheat straw was studied in this paper. Through experiments and analysis, 0.8MPa operation pressure and 1:20 wheat straw to water ratio are optimum for destroying lignin and the maximum of lignin loss rate is 19.94%. After steam explosion pretreatment, the wheat straw was retted by Trametes versicolor for 40 days. Biodegradation rate of lignin was tested and the maximum of 55.40% lignin loss rate was found on day 30. During the whole process of both steam explosion pretreatment and biodegradation, 75.34% lignin was degraded, without steam explosion the biodegradation of raw material the degradation rate of lignin was 31.23% only. FT-IR spectroscopy, TGA and SEM were used for further validating the results of biodegradation.     

Peroxyl radicals promoted changes in water permeability through gramicidin channels in DPPC and lecithin-PC vesicles

Gramicidin incorporation to DPPC or lecithin-PC large unilamellar vesicles (LUVs) leads to pore formation that, under hyper-osmotic conditions, produces a noticeable increase in the rate of trans-membrane water flow. This pore formation is more efficient in the more fluid lecithin-PC LUVs. Exposure of these vesicles to peroxyl radicals generated in the aerobic thermolysis of 2,2'-azo-bis(2-amidinopropane) (AAPH), changes the physical properties of the bilayer (as sensed employing fluorescent probes), modifies gramicidin molecules (as sensed by the decrease in Trp fluorescence) and notably reduces the transbilayer rate of water outflow. In order to evaluate if this reduced water-transport capacity is due to changes in the membrane due to lipid-peroxidation and/or direct damage to gramicidin channels, results obtained in the oxidable vesicles (lecithin-PC) were compared to those obtained in DPPC vesicles. The data obtained show that most of the water transport efficiency loss can be ascribed to a direct disruption of gramicidin channels by AAPH derived peroxyl radicals.     

General method for determining anaerobic biodegradation potential

A simple, generalized method was refined and validated to test whether an organic chemical was susceptible to anaerobic degradation to CH4 + CO2. The method used digested sewage sludge diluted to 10% and incubated anaerobically in 160-ml serum bottles with 50 micrograms of C per ml of test chemical. Biodegradation was determined by the net increase in gas pressure in bottles with test chemicals over the pressure in nonamended sludge bottles. Gas production was measured by gas chromatography and by a pressure transducer. The latter method is recommended because of its speed, accuracy, and low cost. Sewage sludge from municipal digesters with 15- to 30-day retention times was found to be suitable. The sludge could be stored anaerobically at 4 degrees C for up to 4 weeks with satisfactory test results. p-Cresol, phthalic acid, and ethanol are suggested as reference chemicals to confirm sludge activity and method reliability. A revised anaerobic salts medium was developed which minimizes problems of a biological gas production (CO2), avoids precipitation, and meets the requirements of the anaerobic microbiota. When greater than 75% of the theoretical gas production was observed, the chemical was judged to be degradable, and when 30 to 75% of the expected gas was produced, it was termed partially degradable. This method has been tested on more than 100 chemicals of various physical properties and found to reproducibly determine anaerobic biodegradation potential. Of the chemicals tested, 46 were found to be anaerobically degraded. Sludges from nine different municipal treatment plants were surveyed for their ability to degrade nine chemicals which differed in susceptibility to degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

14C-[lignin]-lignocellulose biodegradation by bacteria isolated from polluted soil

Four bacterial species [Branhamella catarrhalis (gram -ve), Brochothrix species (gram -ve), Micrococcus luteus (gram +ve) and Bacillus firmus (gram +ve)], isolated from the soil polluted with cane sugar factory effluents, were found capable of growing on solid media supplemented with indulin AT (a polymeric industrial lignin) as sole C source. All the four species could metabolize cinnamic acid (a non-hydroxylated phenylpropanoid) as sole carbon source with significant suppression on addition of readily metabolizable carbon source (glucose). However, Br. catarrhalis and Brochothrix sp. were capable of metabolizing ferulic acid, but could not do so on addition of glucose. Of the four species, Br. catarrhalis could evolve significant amount of 14CO2 from U-14C (lignin)-lignocellulose prepared from rice stalks (ca. 10% of the added radioactivity in 3 weeks), in addition to solubilization of another 11.7% radioactivity in culture filtrate. The other three species could not significantly evolve 14CO2, though a significant fraction of added 14C-lignin (6.1 to 11.2%) could be solubilized into culture filtrate, suggesting lack of ring-cleavage or other CO2 evolving mechanisms in these species.     

Oxygen radicals and plasma membrane potential

Plasma membrane potential is an important physiologic parameter dependent both upon cellular metabolism and upon the integrity of the cell membrane. Oxygen radicals have been shown to produce significant and early changes in the membrane potential. This data is reviewed along with the physico-chemical origins of the membrane potential and methods of its measurement.     

The influence of lignin content and temperature on the biodegradation of lignocellulose in composting conditions

The aim of this research was to study the influence of lignin content and composting temperature on the biodegradation of lignin-containing pulp and paper products in a controlled composting test (European standard prEN 14046). Lignin reduced the biodegradation of the samples, and there was a linear correlation between the lignin content and the biodegradation of pulp and paper products at 58°C. The influence of incubation temperature (35, 50 and 58°C) on biodegradation was studied using bleached kraft paper containing 0.2 wt% lignin and mechanical pulp (stone-ground wood) containing 24–27 wt% lignin. Mechanical pulp biodegraded better at lower temperatures, while kraft paper biodegraded well at all three temperatures. Microbial activity was evaluated by measuring CO₂ evolution and the change in ATP content, and fungal biomass by measuring the ergosterol content during the composting experiments. Kraft paper strongly increased microbial activity during the controlled composting test, but the activity returned to the background level at the end of the composting test. The proportion of sample carbon converted to microbial biomass carbon was considerably higher at lower incubation temperatures. Changes in microbial community structure during biodegradation of mechanical pulp and kraft paper at 50°C were studied by the PCR-based technique denaturing gradient gel electrophoresis. Changes in the microbial community were observed during the intensive degradation phase of kraft paper. Electronic Publication     

Actinomycetes as agents of biodegradation in the environment--a review.

The diversity of form in the Actinomycetales is well-recognised, due to the sustained generation of environmental isolates for pharmaceutical screening. Actinomycetes isolated from soil and related substrates show primary biodegradative activity, secreting a range of extracellular enzymes and exhibiting the capacity to metabolise recalcitrant molecules. Composting is one process which relies heavily on such prolific actinomycete activity. Amongst actinomycetes in soil, there are examples of different strategies, from cycles of rapid proliferation and sporulation to the maintenance of populations by prolonged slow growth and scavenging, and the evidence for this is examined. The mechanisms of lignocellulose degradation by actinomycetes are discussed in relation to functional conservation within the group, and correlations with those described in other bacteria and fungi.     

Litter decay rates are determined by lignin chemistry

Litter decay rates are often correlated with the initial lignin:N or lignin:cellulose content of litter, suggesting that interactions between lignin and more labile compounds are important controls over litter decomposition. The chemical composition of lignin may influence these interactions, if lignin physically or chemically protects labile components from microbial attack. We tested the effect of lignin chemical composition on litter decay in the field during a year-long litterbag study using the model system Arabidopsis thaliana. Three Arabidopsis plant types were used, including one with high amounts of guaiacyl-type lignin, one with high aldehyde- and p-hydroxyphenyl-type lignin, and a wild type control with high syringyl-type lignin. The high aldehyde litter lost significantly more mass than the other plant types, due to greater losses of cellulose, hemicellulose, and N. Aldehyde-rich lignins and p-hydroxyphenyl-type lignins have low levels of cross-linking between lignins and polysaccharides, supporting the hypothesis that chemical protection of labile polysaccharides and N is a mechanism by which lignin controls total litter decay rates. 2D NMR of litters showed that lignin losses were associated with the ratio of guaiacyl-to-p-hydroxyphenyl units in lignin, because these units polymerize to form different amounts of labile- and recalcitrant-linkages within the lignin polymer. Different controls over lignin decay and polysaccharide and N decay may explain why lignin:N and lignin:cellulose ratios can be better predictors of decay rates than lignin content alone.     

General method for determining anaerobic biodegradation potential.

A simple, generalized method was refined and validated to test whether an organic chemical was susceptible to anaerobic degradation to CH4 + CO2. The method used digested sewage sludge diluted to 10% and incubated anaerobically in 160-ml serum bottles with 50 micrograms of C per ml of test chemical. Biodegradation was determined by the net increase in gas pressure in bottles with test chemicals over the pressure in nonamended sludge bottles. Gas production was measured by gas chromatography and by a pressure transducer. The latter method is recommended because of its speed, accuracy, and low cost. Sewage sludge from municipal digesters with 15- to 30-day retention times was found to be suitable. The sludge could be stored anaerobically at 4 degrees C for up to 4 weeks with satisfactory test results. p-Cresol, phthalic acid, and ethanol are suggested as reference chemicals to confirm sludge activity and method reliability. A revised anaerobic salts medium was developed which minimizes problems of a biological gas production (CO2), avoids precipitation, and meets the requirements of the anaerobic microbiota. When greater than 75% of the theoretical gas production was observed, the chemical was judged to be degradable, and when 30 to 75% of the expected gas was produced, it was termed partially degradable. This method has been tested on more than 100 chemicals of various physical properties and found to reproducibly determine anaerobic biodegradation potential. Of the chemicals tested, 46 were found to be anaerobically degraded. Sludges from nine different municipal treatment plants were surveyed for their ability to degrade nine chemicals which differed in susceptibility to degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization and genomic analysis of kraft lignin biodegradation by the beta-proteobacterium Cupriavidus basilensis B-8

Background

Lignin materials are abundant and among the most important potential sources for biofuel production. Development of an efficient lignin degradation process has considerable potential for the production of a variety of chemicals, including bioethanol. However, lignin degradation using current methods is inefficient. Given their immense environmental adaptability and biochemical versatility, bacterial could be used as a valuable tool for the rapid degradation of lignin. Kraft lignin (KL) is a polymer by-product of the pulp and paper industry resulting from alkaline sulfide treatment of lignocellulose, and it has been widely used for lignin-related studies.

Results

Beta-proteobacterium Cupriavidus basilensis B-8 isolated from erosive bamboo slips displayed substantial KL degradation capability. With initial concentrations of 0.5–6 g L^-1, at least 31.3% KL could be degraded in 7 days. The maximum degradation rate was 44.4% at the initial concentration of 2 g L^-1. The optimum pH and temperature for KL degradation were 7.0 and 30°C, respectively. Manganese peroxidase (MnP) and laccase (Lac) demonstrated their greatest level of activity, 1685.3 U L^-1 and 815.6 U L^-1, at the third and fourth days, respectively. Many small molecule intermediates were formed during the process of KL degradation, as determined using GC-MS analysis. In order to perform metabolic reconstruction of lignin degradation in this bacterium, a draft genome sequence for C. basilensis B-8 was generated. Genomic analysis focused on the catabolic potential of this bacterium against several lignin-derived compounds. These analyses together with sequence comparisons predicted the existence of three major metabolic pathways: β-ketoadipate, phenol degradation, and gentisate pathways.

Conclusion

These results confirmed the capability of C. basilensis B-8 to promote KL degradation. Whole genomic sequencing and systematic analysis of the C. basilensis B-8 genome identified degradation steps and intermediates from this bacterial-mediated KL degradation method. Our findings provide a theoretical basis for research into the mechanisms of lignin degradation as well as a practical basis for biofuel production using lignin materials.     

A colorimetric assay for lignin based on its reaction with diazotized sulfanilic acid and its use in studies on lignin biodegradation by bacteria

Summary Studies on lignin biodegradation are considerably hampered by the lack of a simple analytical method. A rapid colorimetric assay for lignin has been developed using its reaction with a diazotized derivative of sulfanilic acid. Using such a method degradation of lignin, by an isolateErwinia sp. Cu3614, and by a genetically engineeredAcinetobacter PE7(pDPE2388) containing the arylether cleaving gene fromErwinia, has been followed.     

Photochemical, photophysical and photobiological properties of some methylallopsoralens which are potential agents for photochemotherapy

Two new trimethylallopsoralens, 4,7,4'- and 4,7,5'-trimethylallopsoralen, form molecular complexes with DNA and by successive UVA (320-400 nm) irradiation photobind monofunctionally to the macromolecule. The DNA photobinding rates at 365 nm and photobinding quantum yields at 330 nm are markedly higher for 4,7,4'-trimethylallopsoralen than for 4,7,5'-trimethylallopsoralen. Their capacities to generate singlet oxygen in water and benzene are low, particularly for 4,7,4'-trimethylallopsoralen, and the two trimethylallopsoralens completely lack skin phototoxicity on guinea-pig skin. Both compounds show antiproliferative activity in terms of DNA synthesis inhibition in Ehrlich cells and T2 phage infectivity higher than that displayed by angelicin. In view of its monofunctional character, lack of skin phototoxicity, low singlet oxygen yield and antiproliferative activity, 4,7,4'-trimethylallopsoralen deserves further clinical studies as a potential photochemotherapeutic agent.