Highest Manganese Peroxidase Activity Research Articles

Screening of Conditions for Manganese Peroxidase Production from White Rot Fungi (Pleurotus djamor)

The need for alternative and environmentally friendly methods for waste clean-up has led to the use of enzymes in bioremediation. In this study, white rot fungi were isolated from decaying plant parts using standard microbiological and biochemical techniques. The isolated fungi were identified and screened using standard substrates (2, 6-DMP) to determine their capability to produce manganese peroxidase. Pure manganese peroxidase was achieved after four distinct purification phases. Crude extract of the homogenate proteins obtained through optimized submerged fermentation system were precipitated using ammonium sulfate. Ammonium sulfate used at 60% concentration at pH 4.5 precipitated proteins with highest Manganese peroxidase activity (322 U/mg). The precipitated proteins were desalted through dialysis for twelve hours with buffer exchange at interval of six hours and activity of 343.91 U/mg was recorded afterwards. DEAE-cellulose was used for the ion exchange purification of the dialyzed protein, NaCl gradients of 0.3-0.6 M was found to be best in washing off bound protein from the exchange resin and activity of 434.18 U/mg was recorded from the pooled fraction tubes. Sephadex G-100 was used for separation of the proteins into molecular sizes and weights. 2.8 purification folds of the enzyme were achieved after ion exchange (DEAE-cellulose) and gel filtration (sephadex G-100) with percentage yield of 2.20%. Specific activity of the enzyme increased to 602.00% after gel filtration. The partial purified enzyme was further characterized for determination of optimal pH (4.5), temperature (40 OC) and kinetic properties Km of 3.4 mM and Vmax of 250µmol/min.

Using of Oxidoreductase Producing Consortium with High Manganese Peroxidase Activity for Melanoidin Degradation and Electricity Generation from Palm Oil Mill Effluent

Melanoidin is the dark high molecular weight formed via the Maillard reaction under high-temperature conditions. Here, the crude enzyme of melanoidin degrading bacterial consortium was extracted and determined the oxidoreductase activities like manganese peroxidase (MnP), laccase, and lignin peroxidase (LiP). The melanoidin degrading ability and electricity generation were measured. Finally, the oxidoreductase-producing consortium was used as a whole-cell biocatalyst in the air-cathode MFC. The result indicated that the MnP activity of 38.100.10 U/mL was produced from the consortium. The melanoidin removal of 85.102.00% and maximal power output of 475.691.13 W/m3 were achieved. This study gained new knowledge about using oxidoreductase-producing consortium with high MnP activity for melanoidin removal and electricity generation from palm oil mill effluent.

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.

Performance of Meyerozyma caribbica as a novel manganese peroxidase-producing yeast inhabiting wood-feeding termite gut symbionts for azo dye decolorization and detoxification

For hazardous toxic pollutants such as textile wastewater and azo dyes, microbial-based and peroxidase-assisted remediation represents a highly promising and environmentally friendly alternative. Under this scope, gut symbionts of the wood-feeding termites Coptotermes formosanus and Reticulitermes chinenesis were used for the screening of manganese peroxidase (MnP) producing yeasts intended for decolorization and detoxification of textile azo dyes, such as Acid Orange 7 (AO7). To this end, nine out of 38 yeast isolates exhibited high levels of extracellular MnP activity ranging from 23 to 27 U/mL. The isolate PPY-27, which had the highest MnP activity, was able to decolorize various azo dyes with an efficiency ranging from 87.2 to 98.8%. This isolate, which represents the molecularly identified species Meyerozyma caribbica, was successfully characterized in terms of morphological and physiological traits, as well as enzymatic activities. Almost complete decolorization was achieved by the MnP-producing M. caribbica strain SSA1654 after 6 h of incubation with 50 mg/L of the sulfonated azo dye AO7 at 28 °C with an agitation speed of 150 rpm. The maximum decolorization efficiency of AO7 reached 93.8% at 400 mg/L. The decolorization of AO7 was confirmed by Fourier transform infrared (FTIR) and UV–Vis spectral analysis. High performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC–MS) were used to identify AO7 decomposition intermediates. Based on UV–Vis spectra, FTIR, HPLC, and GC–MS analyses, a plausible AO7 biodegradation mechanism pathway was explored, showing azo bond (-N=N-) cleavage and toxic aromatic amines mineralization CO2 and H2O. Microtox® and phytotoxicity assays confirmed that the AO7 metabolites produced by the strain SSA1654 were almost non-toxic compared to the original sulfonated azo dye.

Ligninolytic enzymes production during polycyclic aromatic hydrocarbons degradation: effect of soil pH, soil amendments and fungal co-cultivation.

Soil microorganisms play an important role in the degradation of PAHs and use various metabolic pathways for this process. The effect of soil pH, different soil amendments and the co-cultivation of fungi on the degradation of PAHs in soil and on the activity of ligninolytic enzymes was evaluated. For that purpose, three fungi were studied: Trichoderma viride, Penicillium chrysogenum and Agrocybe aegerita. Biodegradation assays with a mixture of 200ppm PAHs (fluorene, pyrene, chrysene, and benzo[a]pyrene-50ppm each) were set up at room temperature for 8weeks. The maximum laccase activity by solid state fermentation-SSF (7.43 U/g) was obtained by A. aegerita on kiwi peels with 2weeks and the highest manganese peroxidase activity (7.21 U/g) was reached in 4weeks, both at pH 7. Fluorene, pyrene, and benzo[a]pyrene achieved higher degradation rates in soil at pH 5, while chrysene was more degradable at pH 7. About 85-90% of the PAHs were degraded by fungal remediation. The highest degradation of fluorene was achieved by co-cultivation of A. aegerita and P. chrysogenum, remaining 14% undegradable. Around 13% of pyrene stay undegradable by A. aegerita and T. viride and by A. aegerita and P. chrysogenum, both systems supported in kiwi peels, while 11% of chrysene remained in soil by the co-cultivation of these fungi, supported by peanut shells. Regarding benzo[a]pyrene, 13% remained in soil after treatment with A. aegerita. Despite the increase in degradation of some PAHs with co-cultivation, higher enzyme production during degradation was observed when fungi were cultivated alone.

Bioethanol production from rice straw through an enzymatic route mediated by enzymes developed in-house from Aspergillus fumigatus

This study concerns the in-house development of enzymes from Aspergillus fumigatus and their application in separate hydrolysis and fermentation (SHF) processes for bioethanol production from pretreated rice straw. The cellulase and hemicellulase activities of A. fumigatus cultivated in NaOH-pretreated rice straw were much higher than those of A. fumigatus cultivated in raw rice straw. A. fumigatus cultivated in raw rice straw exhibited no lignin-degrading manganese peroxidase (MnP) or laccase enzymatic activity. However, A. fumigatus cultivated in NaOH-pretreated rice straw exhibited high lignin-degrading MnP and laccase enzymatic activity. A. fumigatus completely degraded 1.0% NaOH-pretreated rice straw, yielding 27.89 g/L reducing sugar upon loading of 50 g/L rice straw (based on the original amount of rice straw that went to pretreatment). Saccharification of 1.0% NaOH-pretreated rice straw using 200 FPU/mL crude enzyme released 22.15 g/L reducing sugars in 20 h. The SHF processes resulted in an ethanol concentration and yield of 9.45 g/L and 83.5%, respectively, with Saccharomyces tanninophilus.

Biodeterioration of plasma pretreated LDPE sheets by Pleurotus ostreatus.

Low-density polyethylene (LDPE) waste generates an environmental impact. To achieve the most suitable option for their degradation, it is necessary to implement chemical, physical and biological treatments, as well as combining procedures. Best treatment was prognosticated by Plackett-Burman Experimental Design (PB), evaluating five factors with two levels (0.25 mM or 1.0 gL-1 glucose, 1.0 or 2.0 mM CuSO4, 0.1 or 0.2 mM ABTS [2, 20-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)], pH 4.5 ± 0.2 or 7.0 ± 0.2 and 30 or 90 day incubation), which was reproduced for 150 days. Therefore, PB identified a sequential treatment (plasma followed by fungus) for partial LDPE biodeterioration. Sheets were pretreated with glow discharge plasma (O2, 3.0 x 10−2 mbar, 600 V, 6 min.), followed by Pleurotus ostreatus biodeterioration. Fungus growth, colonization percentage, and pigment generation followed. Laccase (Lac), manganese peroxidase (MnP) and lignin peroxidase (LiP) activities were appraised. Additionally, contact angle (CA), functional group presence and changes and carbonyl and vinyl indices (Fourier transformed infrared spectroscopy) were evaluated. LDPE surface changes were assessed by Young’s modulus, yield strength, scanning electronic microscopy (SEM), Fourier transformed infrared spectroscopy (FTIR) and atomic force microscopy (AFM). Plasma discharge increased hydrophilicity, decreasing CA by 76.57% and increasing surface roughness by 99.81%. P. ostreatus colonization was 88.72% in 150 days in comparison with untreated LDPE (45.55%). After this treatment carbonyl groups (C = O), C = C insaturations, high hydrophilicity CA (16 ± 4) °, and low surface roughness (7 ± 2) nm were observed. However, the highest Lac and LiP activities were detected after 30 days (Lac: 2.817 U Lac g-1 and LiP: 70.755 U LiP g-1). In addition, highest MnP activity was observed at day 120 (1.097 U MnP g-1) only for P. ostreatus treated LDPE. Plasma favored P. ostreatus adsorption, adherence, growth and colonization (88.72%), as well as partial LDPE biodeterioration, supported by increased hydrophilicity and presence of specific functional chemical groups. The approximate 27% changes in LDPE physical properties support its biodeterioration.

Lignin-Degrading Abilities of Novel Autochthonous Fungal Isolates Trametes hirsuta F13 and Stereum gausapatum F28

SUMMARYThe aim of this research is to isolate and identify fungi with high lignin-degrading abilities that are autochthonous to southern Serbian region. Two novel fungal isolates identified as Trametes hirsuta F13 and Stereum gausapatum F28 were selected to assess their ligninolytic enzyme activities and the efficiency of lignin removal from beech wood sawdust. Obtained results show that both isolates are good sources of industrially valuable enzymes with a potential for application in various biotechnological and industrial processes. Both isolates showed laccase, manganese-dependent peroxidase, and versatile peroxidase activities, while only S. gausapatum F28 had lignin peroxidase activity. This is the first record of the ability of S. gausapatum species to produce lignin peroxidase. T. hirsuta F13 showed higher laccase activity than S. gausapatum F28, while S. gausapatum F28 had higher manganese peroxidase activity. Also, T. hirsuta F13 exhibited much higher laccase activity under submerged cultivation conditions than solid-state cultivation conditions, which is rare for fungi. This is important for industrial processes since the submerged fermentation is a dominant technique in industry. The test of the efficiency of lignin removal showed that both isolates are efficient lignin decomposers. After five weeks of incubation on beech wood sawdust, the total lignin losses were 33.84% with T. hirsuta F13 and 28.8% with S. gausapatum F28.

Screening and Production of Manganese Peroxidase from Fusarium sp. on Residue Materials

In this study, we report the manganese peroxidase production ability from a Fusarium sp. strain using an inexpensive medium of agriculture residues of either rice straw or wood chips as carbon source. The highest manganese peroxidase activity on rice straw medium and on wood chips was 1.76 U/mL by day 9 and 1.91 U/mL by day 12, respectively.

Decolorization of Synthetic Textile Dye and Enzymes Production byImproved Strains of Pleurotus Species

In the present investigation the efficiency of three species of white rot fungui - Pleurotus and their improved dikaryons (heterokaryons) was assessed for decolorization of Phenol Red (Phenolsulfonphthalein or PSP) and production of lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase enzymes. All the species of Pleurotus i.e. P. flabellatus, P. ostreatus and P. citrinopileatus decolorized PSP well. However, improved dikaryons Pfo 6X9 and Poc 9X6 decolorized the dye more effectively than three species of Pleurotus. The improved dikaryons also showed higher ligninolytic activity than the parental species. Poc 9X6 showed higher LiP (56.14 U), MnP (615.15 U) and Laccase (715.25 U) activity. In the present work different pH, age and concentration of inoculum and effect of surfactant i.e. sodium dodecyl sulphate (SDS) and Tween-80 were analyzed in order to determine the optimum ones to decolorize maximum concentration of dye. 5 ml of 10 days old culture on pH 5.5 and 0.1% Tween-80 supported maximum decolorization of PSP.

EXTRACELLULAR LIGNINOLYTIC ENZYMES PRODUCTION BY Pleurotus eryngii ON AGROINDUSTRIAL WASTES

Pleurotus eryngii (DC.) Gillet (MCC58) was investigated for its ligninolytic ability to produce laccase (Lac), manganese peroxidase (MnP), aryl alcohol oxidase (AAO), and lignin peroxidase (LiP) enzymes through solid-state fermentation using apricot and pomegranate agroindustrial wastes. The reducing sugar, protein, lignin, and cellulose levels in these were studied. Also, the production of these ligninolytic enzymes was researched over the growth of the microorganism throughout 20 days, and the reducing sugar, protein, and nitrogen levels were recorded during the stationary cultivation at 28 ± 0.5°C. The highest Lac activity was obtained as 1618.5 ± 25 U/L on day 12 of cultivation using apricot. The highest MnP activity was attained as 570.82 ± 15 U/L on day 17 in pomegranate culture and about the same as apricot culture. There were low LiP activities in both cultures. The maximum LiP value detected was 16.13 ± 0.8 U/L in apricot cultures. In addition, AAO activities in both cultures showed similar trends up to day 17 of cultivation, with the highest AAO activity determined as 105.99 ± 6.3 U/L on day 10 in apricot cultures. Decolorization of the azo dye methyl orange was also achieved with produced ligninolytic enzymes by P. eryngii using apricot and pomegranate wastes.

PRODUCTION OF LIGNINOLYTIC ENZYMES BY SOLID-STATE FERMENTATION USING Pleurotus eryngii

Pleurotus eryngii (DC.) Gillet (MCC58) was investigated for its ability to produce various ligninolytic enzymes such as laccase (Lac), manganese peroxidase (MnP), aryl alcohol oxidase (AAO), and lignin peroxidase (LiP) by solid-state fermentation (SSF), which was carried out using a support substrate from the fruit juice industry. The chemical content of grape waste from this industry was studied. Also, the production patterns of these extracellular enzymes were researched during the growth of the organism for a period of 20 days and the protein, reducing sugar, and nitrogen levels were monitored during the stationary cultivation. The highest Lac activity was obtained as 2247.62 ± 75 U/L on day 10 in the presence of 750 µM Mn2+, while the highest MnP activity was attained as 2198.44 ± 65 U/L on day 15 in the presence of 500 µM Mn2+. Decolorization of methyl orange and reactive red 2 azo dyes was also achieved with ligninolytic enzymes, produced in SSF of P. eryngii.

Optimization of physical and nutritional factors for synthesis of lignin degrading enzymes by a novel strain of Trametes vericolor

This paper reports the production of ligninase enzymes by a new strain of Trametes versicolor IBL-04 producing a novel pattern of ligninolytic enzymes with highest MnP activities followed by LiP and laccase. In previous studies Trametes versicolor has been reported to produce higher activities of MnP, followed by laccase and LiP. Lignocellulosic substrates including wheat straw, rice straw, banana stalks, corncobs, corn stover, and sugarcane bagasse were used in solid state fermentation (SSF) for the production of ligninases including peroxidase (LiP), manganese peroxidase (MnP), and laccase by Trametes versicolor IBL-04. Maximum production of MnP (998 U/mL), LiP (620 U/mL), and Laccase (49.7 U/mL) was observed after 5 days in the SSF medium containing 5g rice straw (60% w/w moisture) in still culture SSF. Moisture, pH, temperature, inoculums size, additional carbon and nitrogen sources, and surfactants had a significant influence on ligninase synthesis by the fungus. Production of ligninases was substantially enhanced by optimizing SSF production process. Maximum MnP (1775 U/mL), LiP (1663 U/mL), and laccase (99 U/mL) were produced when rice straw (5g) at 66.6 % moisture (w/w) receiving 5ml inoculum was incubated at pH 4.0 and 30oC in the presence of maltose (1% w/w) as carbon source, urea (0.2% w/w) as nitrogen source and 1mM Tween-80 (0.3 ml) as surfactant.

Novel evidence of cytochrome P450-catalyzed oxidation of phenanthrene in Phanerochaete chrysosporium under ligninolytic conditions

The presence of cytochrome P450 and P450-mediated phenanthrene oxidation in the white rot fungus Phanerochaete chrysosporium under ligninolytic condition was first demonstrated in this study. The carbon monoxide difference spectra indicated induction of P450 (130pmolmg(-1) in the microsomal fraction) by phenanthrene. The microsomal P450 degraded phenanthrene with a NADPH-dependent activity of 0.44±0.02min(-1). One of major detectable metabolites of phenanthrene in the ligninolytic cultures and microsomal fractions was identified as phenanthrene trans-9,10-dihydrodiol. Piperonyl butoxide, a P450 inhibitor which had no effect on manganese peroxidase activity, significantly inhibited phenanthrene degradation and the trans-9,10-dihydrodiol formation in both intact cultures and microsomal fractions. Furthermore, phenanthrene was also efficiently degraded by the extracellular fraction with high manganese peroxidase activity. These results indicate important roles of both manganese peroxidase and cytochrome P450 in phenanthrene metabolism by ligninolytic P. chrysosporium.

Influence of mineral and organic fertilization on soil fungi, enzyme activities and humic substances in a long-term field experiment

Changes in microfungal communities, fungal activities and humic substances (HS) in agricultural soils kept under different fertilization regimes were observed and their causal relationships were investigated in a long-term field experiment. Fertilization did not change the abundance of HS-utilizing microfungi and, except for organic amendment alone, total culturable microfungi were also unaffected by this factor. Organic fertilization increased activities of manganese peroxidase (MnP) and proteinase, but decreased endo-1,4-beta-glucanase activity compared to the corresponding control without organic fertilization. In soils treated with mineral fertilizers, the activities of MnP, endo-1,4-beta-glucanase and proteinase were higher than in control without any mineral treatment. Both the aromaticity of fulvic acid and the molar mass of humic acid was lower in soil with organic fertilization, which may be a result of oxidative degradation mediated by higher MnP activity observed in treatments with organic fertilization.

Laccase and Manganese Peroxidase Activities of Phellinus Robustus and Ganoderma Adspersum Grown on Food Industry Wastes in Submerged Fermentation

Phellinus robustus produced both laccase (700-4,000 U l(-1)) and manganese peroxidase (MnP) (1,000-11,300 U l(-1)) in fermentation of nine food wastes, whereas Ganoderma adspersum produced only laccase (600-34,000 U l(-1)). Glucose provided high laccase and MnP activity of P. robustus but repressed enzyme production by G. adspersum. Ammonium sulphate and ammonium tartrate increased the P. robustus laccase yield (3-fold), whereas the accumulation of MnP was not enhanced by additional nitrogen.

Manganese peroxidases, laccases and oxalic acid from the selective white-rot fungus Physisporinus rivulosus grown on spruce wood chips

The white-rot fungus Physisporinus rivulosus T241i produced manganese peroxidase (MnP), laccase, and oxalic acid when it was grown on spruce ( Picea abies) wood chips. This white-rot basidiomycete degrades lignin selectively and is promising for use in biopulping of softwood. During solid-state cultivation on wood chips the fungus produced oxalic acid (28 μmol/g dry wood) and a high MnP activity (24 nkat/g dry wood), while measured laccase activity was negligible. The prepurified laccase and MnP isoforms were fractionated with anion exchange chromatography. Characterization of the isolated MnP fractions indicated the presence of several MnP isoforms with p I values between 3.4 and 3.9 and molecular masses between 47 and 52 kDa. The four laccase isoforms had p I values between 3.1 and 3.3 and molecular masses between 66 and 68 kDa. The MnP and laccase isoform profiles did not vary during the 4-week cultivation. The N-terminal amino acid sequences of four laccase and two MnP isoenzymes resembled those of the two other selective white-rot fungi applied in biopulping, i.e. Ceriporiopsis subvermispora CZ-3 and the strain IZU-154.

Decolorization of synthetic dyes by solid state cultures of Lentinula (Lentinus) edodes producing manganese peroxidase as the main ligninolytic enzyme

The ability of the white-rot fungus Lentinula (Lentinus) edodes to decolorize several synthetic dyes was investigated using solid state cultures with corn cob as substrate. Cultures, containing amido black, congo red, trypan blue, methyl green, remazol brilliant blue R, methyl violet, ethyl violet and Poly R478 at 200 ppm, were completely decolorized after 18 days of incubation. Partial decolorization was observed in the cultures containing 200 ppm of brilliant cresyl blue and methylene blue. High manganese peroxidase activity (2600 U/g substrate), but very low lignin peroxidase (<10 U/g substrate) and laccase (<16 U/g substrate) activities were detected in the cultures. In vitro, the dye decolorization was markedly decreased by the absence of manganic ions and H2O2. These data suggest that manganese peroxidase appear to be the main responsible for the capability of L. edodes to decolorize synthetic dyes.

Decolorization of synthetic dyes by solid state cultures of Lentinula ( Lentinus) edodes producing manganese peroxidase as the main ligninolytic enzyme

The ability of the white-rot fungus Lentinula ( Lentinus) edodes to decolorize several synthetic dyes was investigated using solid state cultures with corn cob as substrate. Cultures, containing amido black, congo red, trypan blue, methyl green, remazol brilliant blue R, methyl violet, ethyl violet and Poly R478 at 200 ppm, were completely decolorized after 18 days of incubation. Partial decolorization was observed in the cultures containing 200 ppm of brilliant cresyl blue and methylene blue. High manganese peroxidase activity (2600 U/g substrate), but very low lignin peroxidase (<10 U/g substrate) and laccase (<16 U/g substrate) activities were detected in the cultures. In vitro, the dye decolorization was markedly decreased by the absence of manganic ions and H 2O 2. These data suggest that manganese peroxidase appear to be the main responsible for the capability of L. edodes to decolorize synthetic dyes.

A Novel Carrier forPhanerochaete chrysosporiumImmobilization

Phanerochaete chrysosporium BKMF-1767 cells were immobilized on different carriers. The optimum carrier according to adsorbed P. chrysosporium cells number (86.38%) was determined to be polystyrene foam, a novel carrier. The conditions for the immobilization of cells on polystyrene foam were optimized and determined as 50 rpm, 37 degrees C, and 2h. The results show that the adsorption of P. chrysosporium on polystyrene foam follows the Langmuir adsorption isotherm. High manganese peroxidase activity (421 U/L) and dry mass (4.7 g/L) were recovered from the batch mode polystyrene foam solid state fermentation system.