Extracellular Ligninolytic Enzymes Research Articles

Pretreatment and saccharification of corn cobs using partially purified fungal ligninozymes

AbstractCorn cobs consist primarily of a lignocellulosic material comprising hemicellulose, cellulose, and lignin in a crystalline state, which is resistant to microbial saccharification. Bioethanol production from corn cobs has rarely been attempted, especially using chemical pretreatment methods.The present study deals with the production and purification of fungal (Phanerochaete chrysosporium MTCC 787 and Pleurotus florida PAU 22‐01) extracellular ligninolytic enzymes – lignin peroxidase (LiP), manganese peroxidase (MnP), and laccase (Lac) – followed by their utilization for the biological pretreatment of corn cobs along with saccharification using commercial cellulase. Crude LiP, MnP, and Lac demonstrated specific activity of 2.23, 2.1, and 2.63 U/mg, respectively.The one‐step purification of crude enzyme using diethyl amino ethyl (DEAE) cellulose ion exchange chromatography resulted in 11.3, 10.1 and 8.62‐fold purification of LiP, MnP and Lac activity, respectively, with corresponding specific activity of 25.1 U/mg (LiP), 21.2 U/mg (MnP) and 22.7 U/mg (Lac) in the partially purified ligninozymes. Using the latter, biological pretreatment of 2.5 g corn cobs in a reaction volume of 30 mL containing approximately 200 units of Lac, Lip and MnP enzymes (in phosphate buffer, pH 6) resulted in a maximum of 78.4% delignification with a saccharification efficiency of 97.1% using commercial cellulases.

Use of sawdust for production of ligninolytic enzymes by white-rot fungi and pharmaceutical removal.

Use of white-rot fungi for enzyme-based bioremediation of wastewater is of high interest. These fungi produce considerable amounts of extracellular ligninolytic enzymes during solid-state fermentation on lignocellulosic materials such as straw and sawdust. We used pure sawdust colonized by Pleurotus ostreatus, Trametes versicolor, and Ganoderma lucidum for extraction of ligninolytic enzymes in aqueous suspension. Crude enzyme suspensions of the three fungi, with laccase activity range 12-43 U/L and manganese peroxidase activity range 5-55 U/L, were evaluated for degradation of 11 selected pharmaceuticals spiked at environmentally relevant concentrations. Sulfamethoxazole was removed significantly in all treatments. The crude enzyme suspension from P. ostreatus achieved degradation of wider range of pharmaceuticals when the enzyme activity was increased. Brief homogenization of the colonized sawdust was also observed to be favorable, resulting in significant reductions after a short exposure of 5 min. The highest reduction was observed for sulfamethoxazole which was reduced by 84% compared to an autoclaved control without enzyme activity and for trimethoprim which was reduced by 60%. The compounds metoprolol, lidocaine, and venlafaxine were reduced by approximately 30% compared to the control. Overall, this study confirmed the potential of low-cost lignocellulosic material as a substrate for production of enzymes from white-rot fungi. However, monitoring over time in bioreactors revealed a rapid decrease in enzymatic ligninolytic activity.

Construction of CRISPR-Cas9 genome editing platform for white-rot fungus Cerrena unicolor BBP6 and its effects on extracellular ligninolytic enzyme biosynthesis

White-rot fungi (WRF) produce extracellular ligninolytic enzymes such as laccase and manganese-dependent peroxidase (MnP). They have a remarkable potential in degrading a wide range of pollutants including polycyclic aromatic hydrocarbons and synthetic dyes. However, the lack of WRF genetic manipulation methods limits their further applications. Cerrena unicolor BBP6 is a wild-type WRF for laccase and MnP hyper production. This study is the first attempt to develop a genome editing platform for the genus Cerrena sp. based on CRISPR-Cas9 technology. Firstly, the pyrG gene in C. unicolor BBP6 was successfully disrupted and used as an effective screening marker. Subsequently, two genes, LacA and mnp3, which respectively encode a laccase and a MnP protein in strain BBP6, were disrupted. The laccase activity of the LacA mutant strain was reduced by 53.5% compared to the wild-type strain while the MnP activity of the mnp3 mutant strain was reduced by 64.7%. Our study demonstrated that the CRISPR-Cas9 genome-editing platform for C. unicolor BBP6 was successfully established, and it was effective on perturbation of ligninolytic enzyme biosynthesis by this fungus.

Ligninolytic enzymes in Basidiomycetes and their application in xenobiotics degradation

Variety of microorganisms have already proven their capabilities for degradation of wide range of wastes with anthropogenic nature. These pollutants, both liquid and solids, also include so called xenobiotics like phenol and its derivatives, PAHs, dyes, pesticides, pharmaceuticals, etc. Xenobiotics as bisphenol A (BPA), chlorhexidine (CHX), octenidine (OCT), other disinfectants and antiseptics have high ecotoxicological impact. Moreover, they can also impair our quality of life and our health interfering different metabolic and hormone receptors pathways in human body. Chemical treatment of such wastes is not a viable option because of its poor socio-economics and environmental merits. Therefore, applying effective, ecofriendly and cheap treatment methods is of great importance. Basidiomycetes are extensively investigated for their abilities to degrade numerous pollutants and xenobiotics. Through their extracellular ligninolytic enzymes they are capable of reducing or completely removing wide range of hazardous compounds. These enzymes can be categorized in two groups: oxidases (laccase) and peroxidases (manganese peroxidase, lignin peroxidase, versatile peroxidase). Due to the broad substrate specificity of the secreted enzymes Basidiomycetes can be applied as a powerful tool for bioremediation of diverse xenobiotics and recalcitrant compounds.

The synergism of manganese peroxidase and laccase from Cerrena unicolor BBP6 in denim dye decolorization and the construction of gene co-expression system in Pichia pastoris

Extracellular ligninolytic enzymes of white-rot fungi including laccase and manganese peroxidase (MnP), are being applied to denim bleaching process due to their superior degradation capability of synthetic dyes. In the process of denim bleaching by the extracellular broth of Cerrena unicolor BBP6, MnP was primarily responsible for the decolorization, while laccase played an assisting role in the decolorizing reaction by participating in the production of H2O2 to initiate the catalytic reaction of MnP. In our case, the concentration of H2O2 (50 μM) generated from the oxidation reaction of Mn2+ by laccase (2869 U L−1), guaranteed that 200 U L−1 of MnP could carry out its significant decolorization ability of denims without exogenous H2O2 in the bleaching system. A gene co-expression system of Pichia pastoris was then constructed with the highest recombinant MnP and laccase activity of (178.8 and 6390.6, U L−1), respectively. The recombinant enzyme broth on day 6 containing the highest MnP activity and appropriate concentration of H2O2 showed a significantly enhanced decolorization effect on denims. Our results demonstrated that the recombinant enzyme complex (rMnP3-BBP6 and rLaccaseA-189) has shown promising application potential in textile industries, especially in denim bleaching process.

Deciphering the Amendment-Induced Secretion of Ligninolytic Enzymes by Pleurotus pulmonarius

Pleurotus pulmonarius belonging to the white-rot fungal basidiomycetes group secretes extracellular ligninolytic enzymes for the degradation of agroresidues. The present study was carried out to evaluate the efficacy of different agro-residues for the enhanced production of ligninolytic enzymes and to authenticate their ability by protein analysis. The morphological and molecular sequences of white-rot fungi were characterised. Besides, the efficacy of organic and inorganic amendments in the secretion of ligninolytic enzymes by P. pulmonarius was characterised using SDS-PAGE and native PAGE analysis. The characterised strain of P. pulmonarius secreted enhanced laccase enzyme levels in the liquid medium through supplementation with organic and inorganic amendments. Wheat bran and groundnut cake each @5% enhanced secretions of Laccase, LiP and MnP. Copper sulphate at 150 μM enhanced the laccase enzyme and at 100 μM enhanced the LiP enzyme level by P. pulmonarius. Similarly, supplementation with manganese sulphate at 150 μM enhanced laccase, LiP and MnP enzyme levels compared to control. SDS-PAGE results showed protein banding patterns in the range of 50–85 kDa for the Lac enzyme in samples drawn from wheat bran and groundnut cake-supplemented substrates. Native PAGE results of laccase enzymes also showed that wheat bran (5%) + groundnut cake (5%) + CuSO4 (150 M) + MnSO4 (150 M) induced four laccase isozymes. Supplementing organic and inorganic amendments to the substrates would enhance the secretion of laccase enzyme that would aid in better breakdown of lignin.

Effect of Pretreated Colza Straw on the Growth and Extracellular Ligninolytic Enzymes Production by Lentinula edodes and Ganoderma lucidum

Lentinula edodes 3565 and Ganoderma lucidum 9621 were compared for their ability to produce lignocellulolytic enzymes in submerged (SM) and surface liquid (SL) fermentation of hydrolysed colza straw lignin waste that remained after the production of furfural and bioethanol (CS lignin). Application of cultivated mushrooms to dispose of pretreated colza straw agricultural waste is an approach to decrease the quantity of residual lignin while simultaneously obtaining active substances, e.g., the ligninolytic enzyme complex from mycelium. The effect of adding CS lignin to culture media on the yield of L. edodes and G. lucidum mycelium and extracellular laccase activity was studied. It was revealed that the mycelial growth of G. lucidum on solid media was significantly improved by adding CS lignin. Laccase activity during SL cultivation of L. edodes on medium with CS lignin gradually increased over the experiment starting on day 21 and peaked at 520 U/mL on day 28. G. lucidum expressed the maximum laccase activity, 540 U/mL, during the first 14 days of mycelium SM cultivation. Extracellular laccase activity was enhanced about 35- to 40-fold at cultivation of L. edodes and about 10- to 15-fold in the case of G. lucidum by supplementing liquid culture media with CS lignin.

Innovative mycoremediation technique for treating unsterilized PCDD/F-contaminated field soil and the exploration of chlorinated metabolites

Mycoremediation of unsterilized PCDD/F-contaminated field soil was successfully demonstrated by solid-state fermentation coupled with Pleurotus pulmonarius utilizing a patented incubation approach. The experiments were carried out in four setups with two as controls. The contaminated soil was homogenously mixed with solid inocula, 1:0.5 dry w/w, resulting in an initial concentration of 4432 ± 623 ng WHO-TEQ kg−1. After a 30-day incubation under controlled conditions, the overall removal (approx. 60%) was non-specific. The removal was attributed to degradation by extracellular ligninolytic enzymes and uptake into the fruiting tissue (~110 ng WHO-TEQ kg−1 of mushroom). Furthermore, less recalcitrant chlorinated metabolites were found, implying ether bond cleavage and dechlorination happened during the mycoremediation. These metabolites resulted from the complex interaction between P. pulmonarius and the indigenous microbes from the unsterilized soil. This study provides a new step toward scaling up this mycoremediation technique to treat unsterilized PCDD/F-contaminated field soil.

Biopulping Of Bagasse Using Different Types Of White Rot Fungi and Different Incubation Times

Bagasse is fibrous residue that remains after the sugarcane is crushed to extract it’s juice. It mostly consists of lignocellulosic materials that may provide material for paper production through biopulping process. White rot fungi (WRF) is producers of extracellular ligninolytic enzymes that has the capability to mineralize lignin compounds. Three types of white rot fungi that were used in this study are Phanerochaete chrysosporium, Pleurotus ostreatus and Schizophyllum commune. Each of it was inoculated on bagasse substrate within 0, 15, and 30 days of incubation. This study was aimed to know the effect of interaction between white rot fungi and incubation time in the biopulping process and to investigate the most appropriate fungus and incubation time to produce good material for paper making obtained from sugarcane bagasse. The experimental design was done by using Completely Randomized Design (CRD) with a factorial pattern in two factors and analyzed by using Analysis of Variance (ANNOVA) then followed by Duncan’s Multiple Range Test (DMRT) on highly different significance effect of the treatment. The result of this study showed that S. commune is the most effective fungi to degrade highly lignin content (17.38% to 8.88%) at 30 days of incubation, while P. chrysosporium is the most effective fungi to lowering cellulose content in small amount (23.64% to 19.38%) during 30 days of incubation.

Studies on cultivation, nutrition and extracellular ligninolytic enzymes of Flammulina velutipes strains collected from Indian Himalayas

Seven different strains of Flammulina velutipes collected from the North East and North West Himalayas were evaluated for total protein and ligninolytic enzymes on wheat straw and malt extract liquid broth medium respectively. Nutritional analyses of 6 strains were also done followed with yield evaluation. Maximum mycelia protein was observed in strain DMRX-166 on the 20th day in malt extract medium and DMRX-768 and DMRX-769 in wheat straw medium. Minimum laccase production was observed in strain (DMRO-253). Laccase appearance starts on the 5 day of incubation and continuously increased upto 20th37th day. Highest peroxidase activity was observed in DMRX-897 in wheatstraw medium on 15 day followed by DMRO-253. In general, more peroxidase activity observed in wheat straw media as compared to malt extracts media. On the 15th day strain no. DMRX-1618 produced maximum ligninase activity (175 U/ml) followed by DMRX-1446 on malt extract medium. Wheat straw broth medium favored early lignin peroxidase activity on the 10th day in 3 strains while 4 strains produce more lignin peroxidase on the 20th day. Least lignin peroxidase activity was recorded inDMRX-897 in the malt extract medium. Manganese peroxidase was observed in DMRO-253 in malt extract medium. Nutritional analysis showed highest protein content in DMRX-1446 while highest vitaminC and D2th were found in DMRX-767. The current study elucidated enzyme and nutritional relationship with time will help for developing efficient cultivation practices in Flammulina in near future.

A proposed stepwise screening framework for the selection of polycyclic aromatic hydrocarbon (PAH)-degrading white rot fungi.

This study suggests a simple three-step screening protocol for the selection of white rot fungi (WRF) capable of degrading polycyclic aromatic hydrocarbons (PAHs), which combines easily applicable bioassay techniques, and verifies that protocol by evaluating the PAH degradation activity, ligninolytic enzyme secretion, and relevant gene expressions of the selected PAH-degraders. Using 120 fungal strains, a sequence of bioassay techniques was applied: Bavendamm's reaction (Step 1), remazol brilliant blueR (RBBR) decolorization (Step 2); assays for tolerance to four mixed PAHs-phenanthrene, anthracene, fluoranthene, and pyrene (Step 3). This stepwise protocol selected 14 PAH-degrading WRF, including Microporus vernicipes, Peniophora incarnata, Perenniporia subacida, Phanerochaete sordida, Phlebia acerina, and Phlebia radiata. Of these, P. incarnata exhibited the highest PAH degradative activity, ranging from 40 to > 90%, which was related to the time-variable secretions of three extracellular ligninolytic enzymes: laccase, manganese-dependent peroxidase (MnP) and lignin peroxidase (LiP). Laccase and MnP production by P. incarnata tended to be greater in the early stages of PAH degradation, whereas its LiP production became intensified with decreasing laccase and MnP production. Pilc1 and pimp1 genes encoding laccase and MnP were expressed, indicating the occurrence of extracellular enzyme-driven biodegradation of PAH by the fungal strains.

Enhancement of laccase production in a new isolated Trametes hirsuta LBF-AA017 by lignocellulosic materials and its application for removal of chemical dyes

Laccase is one of the extracellular ligninolytic enzymes highly produced by white rot fungi (WRF) and widely used in industrial processes such as decolorizing of synthetic dyes. In this study, the enzyme was produced by a new isolated WRF (Trametes hirsuta LBF-AA017) through two types of fermentation: submerged and solid state fermentation. In the submerged culture, several local lignocellulosic materials (0.5% w/v): bagasse, palm kernel cake (PKC), sugar palm fruit cake (SPFC), corn cobs (CC), tobacco road (TR), were used as laccase inducers. Among 5 materials tested, treatment using SPFC resulted highest laccase production, 645 U L−1 in the 9th day. Higher concentration of SPFC (5% w/v) enhanced laccase poduction by the fungus up to 2034.354 U L−1 in 9 days. Addition of SPFC above 25% (w/v) into culture medium made solid state condition and significantly enhance the production of laccase by the fungus. The crude laccase could effectively decolorize three types of chemical dyes: congo red (CR), brilliant blue G (BBG), coomassie brilliant blue (CBB), in a short period. The laccase could effectively remove three kind of tested dyes with various rates: 45, 91, 48%, respectively for CR, BBG and CBB in one hour reaction. Rapid removal of dyes, especially for BBG, by the laccase T. hirsuta LBF-AA017 has initially proven the potential to be applied for environmental biotechnology.

Mycoremediation of 17 β‐Estradiol Using Trichoderma citrinoviride Strain AJAC3 along with Enzyme Studies

Extensive and inappropriate use of steroids in drugs and pharmaceutical products and improper release of steroids into the immediate environment has found to affect soil and water resulting in imbalance of ecosystem. The steroids reach environment due to inaccurate disposal of expired drugs as well as manufacturing disposal from pharmaceutical industries and are urgently required to be removed from the environment. The present study deals with elimination of 17 β‐estradiol (E2) using Trichoderma citrinoviride AJAC3. The fungal isolate could tolerate E2 and degrade 99.6% of the contaminant at concentration of 200 mg L−1. Trichoderma citrinoviride AJAC3 secreted extracellular ligninolytic enzymes which played significant role in degradation of E2. High Performance Liquid Chromatography (HPLC) was used to determine the biodegradation pattern of E2 and confirmed by Gas Chromatography Mass Spectroscopy (GC–MS), Scanning Electron Microscopy (SEM), and Fourier Transform Infrared Spectroscopy (FTIR) analysis. Kinetics study revealed that the degradation pathway followed first‐order kinetic model. The results affirmed that the isolate was able to biodegrade E2 efficiently. © 2019 American Institute of Chemical Engineers Environ Prog, 38:e13142, 2019

Production of Ligninolytic Enzymes by Coptotermes curvignathus Gut Bacteria

Abstract Maximum utilization of lignocellulosic biomass is contingent upon degrading the recalcitrant lignin polymer. Conventional methods employed in delignification require high inputs of energy and chemicals, resulting in the release of highly toxic effluents. The ability of gut flora of Coptotermes curvignathus in lignin degradation was investigated in this study. Production of ligninolytic enzymes was done in an aerated submerged fermentation system with kraft lignin as sole carbon source. The degradation experiment was carried out for 7 days at 30 °C, pH 7. Three potential lignin degraders identified as Bacillus sp., Lysinibacillus sp. and Acinetobacter sp. were successfully isolated. The bacterial growth and secretion of extracellular ligninolytic enzymes confirmed metabolism of kraft lignin by the identified strains. Lysinibacillus sp., a novel lignin degrader showed highest manganese peroxidase (76.36 ± 15.74 U/L) and laccase activity (70.67 ± 16.82 U/L) after 7 and 6 days of incubation respectively, while maximal activity of lignin peroxidase (262.49 ± 0.92 U/L) was recorded after 7 days in culture supernatants of Bacillus sp. With respect to the activity of the secreted enzymes, the lignin degrading potential of these bacterial strains can be explored in the valorisations of lignocellulosic biomass in industrial processes such as pulping, bioethanol production, fine chemicals and materials synthesis.

Dye-decolorizing peroxidases in Irpex lacteus combining the catalytic properties of heme peroxidases and laccase play important roles in ligninolytic system

BackgroundThe white rot fungus Irpex lacteus exhibits a great potential in biopretreatment of lignocellulose as well as in biodegradation of xenobiotic compounds by extracellular ligninolytic enzymes. Among these enzymes, the possible involvement of dye-decolorizing peroxidase (DyP) in lignin degradation is not clear yet.ResultsBased on the extracellular enzyme activities and secretome analysis, I. lacteus CD2 produced DyPs as the main ligninolytic enzymes when grown in Kirk’s medium supplemented with lignin. Further transcriptome analysis revealed that induced transcription of genes encoding DyPs was accompanied by the increased expression of transcripts for H2O2-generating enzymes such as alcohol oxidase, pyranose 2-oxidase, and glyoxal oxidases. Meanwhile, accumulation of transcripts for glycoside hydrolase and protease was observed, in agreement with abundant proteins. Moreover, the biochemical analysis of IlDyP2 and IlDyP1 confirmed that DyPs were able to catalyze the oxidation of typical peroxidases substrates ABTS, phenolic lignin compounds DMP, and guaiacol as well as non-phenolic lignin compound, veratryl alcohol. More importantly, IlDyP1 enhanced catalytic activity for veratryl alcohol oxidation in the presence of mediator 1-hydroxybenzotriazole, which was similar to the laccase/1-hydroxybenzotriazole system.ConclusionsThe results proved for the first time that DyPs depolymerized lignin individually, combining catalytic features of different peroxidases on the functional level. Therefore, DyPs may be considered an important part of ligninolytic system in wood-decaying fungi.

Application of ligninolytic enzyme of Lentinus polychrous on synthetic dye decolorization

Contamination of water resources by synthetic dyes causes many environmental and health problems. Fungal ligninolytic enzymes have been applied for dye decolorization due to its ability in degrading a wide variety of recalcitrant substances. Extracellular ligninolytic enzyme production by white rot fungus, Lentinus polychrous, grown in glucose containing medium supplemented with rice straw powder and soybean pomace was monitored. Optimum condition for remazol brilliant blue R (RBBR) decolorization was studied by varying initial RBBR concentrations, pH values, and initial crude enzyme concentrations. The result revealed that the ligninolytic enzyme dominantly produced was laccase with an activity of 0.095 U/ml on day 15, and a small amount of manganese peroxidase was also detected. The crude laccase produced from L. polychrous, effectively decolorized the dye within a short period of time, that was approximately 50% of 20 mg/l RBBR was decolorized within the first 2 hours. The optimal pH for RBBR decolorization was 3.0 which had an efficiency of 87% within 6 hours after incubation. Moreover, the best redox mediator of laccase was CuSO4, whose decolorization efficiency was over twice than that of samples without this mediator. The decolorization intensified with increase of CuSO4 concentration but higher concentrations of chromium tended to suppress decolorization.

Investigation of the ability of immobilized cells to different carriers in removal of selected dye and characterization of environmentally friendly laccase of Morchella esculenta

The dyestuffs used in the textile sector are among the major pollutants and present as a polluting agent in the waters poured to the environment. Therefore, synthetic dyes led to the serious environmental pollution and threats to human health. The process of color removal from waste water of the textile industry is gaining importance from ecological point of view. Nowadays, the removal of dye materials is carried out by physical and chemical methods. However, the high cost of these methods and the resulting large amount of concentrated slurry causes problems. In recent years, it has been shown that white rot fungi have the ability to reduce the synthetic dye materials with their extracellular ligninolytic enzymes. Fungal biosorption is an economical, easy and efficient method for removing organic contaminants from aqueous media. In this study; the white rot fungus M. esculenta was immobilized into five different support materials (gelatin, polyurethane, kaolin, cellulose) and the removal of Reactive Orange 16 dye with these immobilized cells was investigated and compared. The best biological degradation of the dye was obtained by immobilization into polyurethane support material with M. esculenta (dye concentration: 10 mgL−1) 90,38%. Samples were analyzed in FT-IR to determine possible metabolites that can be occurred as a result of biodegradation. The M. esculenta laccase, which may be responsible for biodegradation, was partially purified and identified by SDS-PAGE.

Interconnection of Key Microbial Functional Genes for Enhanced Benzo[a]pyrene Biodegradation in Sediments by Microbial Electrochemistry.

Sediment microbial fuel cells (SMFCs) can stimulate the degradation of polycyclic aromatic hydrocarbons in sediments, but the mechanism of this process is poorly understood at the microbial functional gene level. Here, the use of SMFC resulted in 92% benzo[a]pyrene (BaP) removal over 970 days relative to 54% in the controls. Sediment functions, microbial community structure, and network interactions were dramatically altered by the SMFC employment. Functional gene analysis showed that c-type cytochrome genes for electron transfer, aromatic degradation genes, and extracellular ligninolytic enzymes involved in lignin degradation were significantly enriched in bulk sediments during SMFC operation. Correspondingly, chemical analysis of the system showed that these genetic changes resulted in increases in the levels of easily oxidizable organic carbon and humic acids which may have resulted in increased BaP bioavailability and increased degradation rates. Tracking microbial functional genes and corresponding organic matter responses should aid mechanistic understanding of BaP enhanced biodegradation by microbial electrochemistry and development of sustainable bioremediation strategies.

Biodecolourisation and biodegradation of leather dyes by a native isolate of Trametes villosa

Abstract Dyeing is an important step in the leather manufacture process. Effluent from this stage contains some types of synthetic dye that may be a threat to the environment and human health. Biological treatment of dye-containing wastewaters by microorganisms has been presented as a cost effective and promising environmentally friendly alternative. In the present work, the potential of Brazilian native white-rot fungi strains, collected and screened to produce extracellular ligninolytic enzymes, was evaluated for the biodecolourisation and biodegradation of different azo tannery dyes. The strain SCS-10 showed high activity of ligninolytic enzymes and allowed the colour removal of dyes in solid media. This isolate was characterised morphologically and identified as Trametes villosa , based on a molecular analysis of the internal transcribed spacer (ITS) region sequences. T. villosa SCS-10 showed high biodecolourisation efficiency for the dyes assessed, achieving 95.71 ± 1.29, 92.76 ± 0.99 and 96.84 ± 1.39% for Acid Red 357, Acid Black 210 and Acid Blue 161, respectively, at 100 mg L −1 , 30 °C, pH 5.5 and 150 rpm, within 168 h of treatment. Remarkable peaks of laccase activity (1150–1550 U L −1 ) were observed during specific periods in the biodecolourisation process. The complete inhibition of Lac activity by sodium azide (NaN 3 , 0.1 mM) led to biodecolourisation values of 13.29 ± 0.93, 12.30 ± 0.46 and 20.05 ± 2.08% for AR357, AB210 and AB161, respectively. These results confirmed the main role of laccase in colour removal, although biosorption also had a minor involvement in biodecolourisation. In vitro assays also showed the efficiency of decolourisation of the leather dyes. The enzymatic crude extract produced by T. villosa allowed 85.45 ± 3.43 (AR357), 76.96 ± 1.39 (AB210) and 90.17 ± 0.97% (AB161) of biodecolourisation when enhanced by the use of the redox mediator 1-hydroxybenzotriazol (HBT, 1 mM). UV–vis and FTIR spectral analyses confirmed the occurrence of enzymatic biodegradation as the mechanism responsible for colour removal. T. villosa SCS-10 was able to tolerate high concentrations of the dyes (200–1000 mg L −1 ) and a wide range of pH (4.0–8.0) during biodecolourisation. The native isolate T. villosa SCS-10 is considered a suitable candidate for the treatment of dye-polluted wastewater from the leather industry due to the mechanisms of enzymatic biodegradation and biosorption.

Decolorization pathways of anthraquinone dye Disperse Blue 2BLN by Aspergillus sp. XJ-2 CGMCC12963

ABSTRACT Anthraquinone dye represents an important group of recalcitrant pollutants in dye wastewater. Aspergillus sp XJ-2 CGMCC12963 showed broad-spectrum decolorization ability, which could efficiently decolorize and degrade various anthraquinone dyes (50 mg L−1) under microaerophilic condition. And the decolorization rate of 93.3% was achieved at 120 h with Disperse Blue 2BLN (the target dye). Intermediates of degradation were detected by FTIR and GC-MS, which revealed the cleavage of anthraquinone chromophoric group and partial mineralization of target dye. In addition, extracellular manganese peroxidase showed the most closely related to the increasing of decolorization rate and biomass among intracellular and extracellular ligninolytic enzymes. Given these results, 2 possible degraded pathways of target dye by Aspergillus sp XJ-2 CGMCC12963 were proposed first in this work. The degradation of Disperse Blue 2BLN and broad spectrum decolorization ability provided the potential for Aspergillus sp XJ-2 CGMCC12963 in the treatment of wastewater containing anthraquinone dyes.