Enhanced biodecolorization of synthetic textile dye effluent by Phanerochaete chrysosporium under improved culture conditions

Biodecolorization of a synthetic commercial textile dye effluent (0.1 gl^–1) by Phanerochaete chrysosporium was enhanced by improving the original Kirk's medium with respect to buffer, C:N ratio, Mg²⁺ and Zn²⁺ ions, inoculum, temperature shifts, agitation and sunflower oil additon. An increase of 6.7-fold in lignin peroxidase (LIP) level, 4-fold in biomass and 45.5% enhanced decolorization of effluent was achieved. Degradation was both enzymatic (47.2%) and by biosorption (61.67%).     

The importance of the morphology and hydrophobicity of different carriers on the immobilization and sugar refinery effluent degradation activity of Phanerochaete chrysosporium

There was no direct correlation between the surface hydrophobicity or hydrophilicity of four solid carriers and the amount of immobilized Phanerochaete chrysosporium. The immobilized biomass was 1.5–1.8 times higher and the fungal degradation activity was 5–8 and 3 times greater in terms of decolorization and phenolics reduction, respectively, with porous carriers than with non-porous carriers. Morphology of the carriers was important and governed the amount of immobilized mycelium and specially the fungal biodegradation activity.     

Use of a fluorescence labelled,carbohydrate-binding module from Phanerochaete chrysosporium Cel7D for studying wood cell wall ultrastructure

The -amino group of the carbohydrate-binding module (CBM) from Phanerochaete chrysosporium cellulase Cel7D was covalently labelled with fluorescein isothiocyanate. The fluorescein-labelled CBM was characterised regarding substrate binding, showing specificity only to cellulose and not to mannan and xylan. Conjugation of fluorescein isothiocyanate to CBM did not affect its binding to cellulose. The labelled CBM was successfully used as a probe for detecting cellulose in lignocellulose material such as never dried spruce and birch wood as well as pulp fibres.     

Kinetics of endosulfan degradation by Phanerochaete chrysosporium

The chlorinated pesticide, endosulfan, could be degraded by Phanerochaete chrysosporium under non-ligninolytic conditions, and this did not require direct contact with mycelium. The major metabolites formed were endosulfan sulfate and endosulfan diol. The rate of degradation depended on the initial concentration. With 2.5 mg endosulfan l^–1, degradation was at 0.23 mg l^–1 day^–1. The degradation could be described using a nonlinear rate expression that was similar to the Michaelis–Menten equation.     

Vanillate hydroxylase from the white rot basidiomycete Phanerochaete chrysosporium

A soluble enzyme fraction from Phanerochaete chrysosporium catalyzed the oxidative decarboxylation of vanillic acid to methoxy-p-hydroquinone. The enzyme, partially purified by ammonium sulfate precipitation, required NADPH and molecular oxygen for activity. NADH was not effective. Optimal activity was displayed between pH 7.5–8.5. Neither EDTA, KCN, NaN₃, nor o-phenanthroline (5 mM) were inhibitory. The enzyme was inducible with maximal activity displayed after incubation of previously grown cells with 0.1% vanillate for 30h.Abbreviations MHQ Methoxy-p-hydroquinone - GLC gas liquid chromatography - TMSi trimethylsilane - TLC thin layer chromatography     

A scalable membrane gradostat reactor for enzyme production using Phanerochaete chrysosporium

This is the first demonstration of process scale-up of a membrane gradostat reactor for continuous enzyme production using Phanerochaete chrysosporium ME446. The fungus was immobilised by reverse filtration on to externally unskinned, ultrafiltration capillary membranes and then nutrient gradients were induced across the biofilm. A 10-fold scale-up from a single capillary bioreactor to a 2.4 l multi-capillary unit resulted in a 7-fold increase in enzyme productivity with a peak at 209 U l^–1 d^–1. Subsequent scale effects on the spore distribution, continuous manganese peroxidase production profile and biofilm development are discussed.     

Secretome analysis of Phanerochaete chrysosporium strain CIRM-BRFM41 grown on softwood

Proteomic analysis was performed to determine and differentiate the composition of the secretomes of Phanerochaete chrysosporium CIRM-BRFM41, a peroxidase hypersecretory strain grown under ligninolytic conditions and on softwood chips under biopulping conditions. Extracellular proteins from both cultures were analyzed by bidimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry. A total of 37 spots were identified. The secretome in liquid synthetic medium comprised mainly peroxidases, while several wood-degrading enzymes and enzymes involved in fungal metabolism were detected in biopulping cultures on softwood. This prompted an analysis of the impact of secretome modulation in the presence of softwood chips. Biotreated wood was submitted to kraft cooking and chemical bleaching using chlorine dioxide. The fungal pre-treatment led to a significant increase in pulp yield and a better bleachability of the pulp. This bleachability improvement could be explained by the production of specific lignocellulose-degrading enzymes.     

Use of Phanerochaete chrysosporium Immobilized on Kissiris for Synthetic Dye Decolourization: Involvement of Manganese Peroxidase

The mineral Kissiris, which is formed from the thickened foam of volcanic lava, was tested to approximate its mineral composition using energy-dispersive X-ray (EDX) analysis. The solid mineral contains silicon dioxide at about 16 (w/w). The considerable surface roughness of Kissiris along with its extensive porosity made this natural solid cell support an attractive candidate for manganese peroxidase (MnP) production for synthetic dye decolourization, at low cost. The white rot fungus Phanerochaete chrysosporium immobilized on the mineral Kissiris was grown in both stationary and agitated cultures (rotary shaker, 100 rev/min) using either carbon- or nitrogen-limited growth medium to study the ability of the fungus to degrade the synthetic dye methylene blue (MB). The value of residual dye for MB used at 60 ppm was 6% within 8 days of the incubation of the nitrogen-limited culture under the shaken conditions. Production of (MnP) occurred simultaneously in nitrogen-limited culture medium with the added MnSO₄ at 100 ppm. The MnP activity was at relatively high level (170 U/l).     

Phenolic removal in olive oil mill wastewater using loofah-immobilized Phanerochaete chrysosporium

Summary Olive oil mill wastewater (OMW) has a high organic load, and this is a serious concern of the olive industry. Conventional biological wastewater treatments, despite their simplicity and suitable performance are ineffective for OMW treatment since phenolics possess antimicrobial activity. In order to carry out a proper treatment of OMW, use of a microorganism able to degrade the phenolics is thus necessary. In this study the ability of Phanerochaete chrysosporium to degrade the phenolic compounds of OMW and to decrease the chemical oxygen demand (COD) using cells immobilized on loofah was examined. The basal mineral salt solution along with glucose, ammonium sulfate and yeast extract was used to dilute the OMW appropriately. The fungus did not grow on the concentrated OMW. The extent of removal in this bio-treatment, of total phenols (TP) and the COD were 90 and 50%, respectively, while the color and aromaticity decreased by 60 and 95%, respectively. The kinetic behavior of the loofah-immobilized fungus was found to follow the Monod equation. The maximum growth rate μ_max was 0.045 h⁻¹ while the Monod constant based on the consumed TP and COD were (mg/l) 370 and 6900, respectively.     

Heterologous expression of lignin peroxidase of Phanerochaete chrysosporium in Pichia methanolica

The cDNA encoding for lignin peroxidase of Phanerochaete chrysosporium was expressed in the Pichia methanolica under the control of the alcohol oxidase (AUG1) promoter which was followed by either the lignin peroxidase leader peptide of Phanerochaete chrysosporium or the Saccharomyces cerevisiae alpha-factor signal peptide. Both peptides efficiently directed the secretion of lignin peroxidase from the recombinant yeast cell. The extracellular lignin peroxidase activity in two recombinants was 932 U l(-1) and 1933 U l(-1). The purity of the recombinant product was confirmed by SDS-PAGE.     

Extracellular ligninolytic enzyme production by Phanerochaete chrysosporium in a new solid-state bioreactor

The production of manganese-dependent peroxidase (MnP) and lignin peroxidase (LiP) by the fungus Phanerochaete chrysosporium (ATCC 24725) in a new bioreactor, the Immersion Bioreactor, which grows cells under solid-state conditions, was studied. Maximum MnP and LiP activities were 987 U l^–1 and 356 U l^–1, respectively. The polymeric dye, Poly R-478, was degraded at 2.4 mg l^–1 min^–1 using the extracellular culture filtrate.     

Production and characterization of cellulolytic and xylanolytic enzymes from the ligninolytic white-rot fungus Phanerochaete chrysosporium grown on sugarcane bagasse

The ligninolytic white-rot fungus Phanerochaete chrysosporium BKM-F-1767 produced extracellular cellulolytic enzymes (carboxymethylcellulase, CMCase and -glucosidase) and xylanolytic enzymes (xylanase and -xylosidase) in liquid medium containing 1.0% sugarcane bagasse with or without 1.0% glucose. The changes in pH and soluble protein content were monitored in the culture filtrates. The results obtained showed that the pH decreased after 3 days and then increased. The soluble protein content increased and reached the maximum value after 12 days. The results showed that the activities of enzymes were higher in the case of sugarcane bagasse without glucose. The characterization study indicated that the optimum pH values were 4.6, 4.2, 5.0 and 5.0 for CMCase, -glucosidase, xylanase and -xylosidase, respectively and the optimum temperatures were 60, 70, 65 and 60 °C for the investigated enzymes, respectively. The results showed also that after prolonged heating (5 h) at 60 °C, CMCase, -glucosidase, xylanase and -xylosidase retained 81.2, 86.8, 51.5 and 27.4% activity, respectively.     

Degradation of pentachlorophenol by the white rot fungus Phanerochaete chrysosporium grown in ammonium lignosulphonate media

Removal and degradation of pentachlorophenol (PCP) by Phanerochaete chrysosporium in static flask cultures was studied using ammonium lignosulphonates (LS), a waste product of the papermill industry, as a carbon and nitrogen source. After 3 days, cultures of P. chrysosporium grown in either a 2% LS (nitrogen-sufficient) medium or a 0.23% LS and 2% glucose (nitrogen-deficient) medium removed 72 to 75% of PCP, slightly less than the 95% removal seen using nitrogen-deficient glucose and ammonia medium. PCP dehalogenation occurred despite the fact that extracellular enzyme (LiP) activity, measured by a veratryl alcohol oxidation assay and by PCP disappearance in cell-free extracts, was inhibited by LS. This inactivation of LiP likely contributed to the lower percent of PCP dehalogenation observed using the LS media. In order to better understand the relationship between PCP disappearance and dehalogenation, we measured the fate of the chlorine in PCP. After 13 days, only 1.8% of the initial PCP added was recoverable as PCP. The remainder of the PCP was either mineralized or transformed to breakdown intermediates collectively identified as organic halides. The largest fraction of the original chlorine (58%) was recovered as organic (non-PCP) halide, most of which (73%) was associated with the cell mass. Of the remaining chlorine, 40% was released as chloride ion, indicating a level of dehalogenation in agreement with previously reported values.     

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     

Utilisation of lignocellulosic wastes for lignin peroxidase production by semi-solid-state cultures of Phanerochaete chrysosporium

In the present work, the production of ligninolytic enzymes by semi-solid-statecultures of Phanerochaete chrysosporium BKM-F-1767 (ATCC 24725),employing different lignocellulosic wastes as support, was investigated. Thewaste materials employed were grape seeds, wheat straw and wood shavings.Maximum lignin peroxidase activities of 1620 ± 123 U/l, 364 ± 35 U/l and 571 ± 42 U/l were attained, respectively. Nevertheless, lowmanganese-dependent peroxidase activities were found, being insignificantin the grape seed cultures. Moreover, the in vivo decolourisation of a model dye compound, the polymeric dye Poly R-478 (polyvinylamine sulfonateanthrapyridone), by the above-mentioned cultures was monitored to assessthe degrading capability of the extracellular liquid secreted by such cultures.The percentage of biological decolourisation attained by grape seed and woodshaving cultures was around 74% and 63%, respectively, whereas it was ratherlow (40%) in the wheat straw ones.     

Biodegradation of bioaccessible textile azo dyes by Phanerochaete chrysosporium

Azo dyes are important chemical pollutants of industrial origin. Textile azo dyes with bioaccessible groups for lignin degrading fungi, such as 2-methoxyphenol (guaiacol) and 2,6-dimethoxyphenol (syringol), were synthesised using different aminobenzoic and aminosulphonic acids as diazo components. The inocula of the best biodegradation assays were obtained from a pre-growth medium (PAM), containing one of the synthesised dyes. The results of the dye biodegradation assays were evaluated every 7 days, by the decrease of the absorbance at the maximum wavelength of the dye, by the decrease of the sucrose concentration in the culture medium and by the increase of the biomass during the 28 days of assay. It was observed that the extent of dye biodegradation depended on the sucrose concentration, on the degraded dye structure and, on the dye present in the PAM medium.     

Remediation of pentachlorophenol-contaminated soil by composting with immobilized Phanerochaete chrysosporium

Summary To reduce and eliminate the hazards of pentachlorophenol (PCP) to the soil, the method of inoculating free and immobilized white rot fungi, Phanerochaete chrysosporium to PCP-polluted soils was investigated. Three parallel beakers A, B, C are adopted with the same components of soil, yard waste, straw and bran for aerated composting to degrade the PCP in soil. A was with no inoculants as control, B was added with the inoculants of immobilized P.␣chrysosporium, C was inoculated with non-immobilized P. chrysosporium, and additionally D contained only PCP-contaminated soils also as control. By contrastive analyses, the feasibility of applying composting to the bioremediation of the PCP-polluted soil was discussed. From the experimental results, it could be seen that the degradation rate of PCP by the immobilized fungi exceeded 50% at day 9, while that of the non-immobilized fungi achieved the same rate at day 16. However, the final degradation rates of PCP for both of them were beyond 90% at day 60 and that the rate of A was much lower than the others. The above data have shown that the degradation effect of inoculating P. chrysosporium was better than that of no inoculation, and that of the immobilized fungi was better than that of non-immobilized ones. Meanwhile, shown by all the indicators the composts of A, B and C were mature and stabilized at the end of the experiment. Therefore, the method of composting with immobilized P.␣chrysosporium is effective for the bioremediation of PCP-contaminated soil.     

Amelioration of ligninolytic enzyme production by Phanerochaete chrysosporium in airlift bioreactors

Maximum activities of manganese-dependent peroxidase (MnP) and lignin peroxidase (LiP) in free cultures of Phanerochaete chrysosporium (ATCC 24725) were 258 U l^–1 and 103 U l^–1, respectively, in an airlift bioreactor. Immobilisation of the fungus on an inert carrier as well as several design modifications of the bioreactor employed gave MnP activities around 500–600 U l^–1 during 9 days' operation. The continuous operation of the latter led to MnP and LiP activities about 140 U l^–1 and 100 U l^–1, respectively, for two months, without operational problems. Furthermore, the extracellular liquid secreted decolourised the polymeric dye Poly R-478 about 56%.     

Protoplast Fusion between Geotrichum candidium and Phanerochaete chrysosporium to Produce Fusants for Corn Stover Fermentation

Lignin impedes the digestion of corn stover when used as an animal feed. Phanerochaete chrysosporium is an efficient lignindegrader. Geotrichum candidum can be used to produce single-cell protein. In this study, protoplasts of the two fungi were prepared and fused. After screening, one of the fusants, Fusant R1, was selected for corn stover fermentation. It decreased lignin from 109 to 54 g/kg and increased protein from 48 to 67 g/kg in corn stover. Comparison with their parental strains indicated that the fusant obtained the lignin-degrading ability from P. chrysosporium and the protein-accumulating ability from G. candidium.