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1.  Decolorization and detoxification of sulfonated toxic diazo dye C.I. Direct Red 81 by Enterococcus faecalis YZ 66 
Isolated Enterococcus faecalis YZ 66 strain shows ability to decolorize various industrial dyes among which, it showed complete decolorization and degradation of toxic, sulfonated recalcitrant diazo dye Direct Red 81 (50 mg/L) within 1.5 h of incubation under static anoxic condition. The optimum pH and temperature for decolorization was 7.0 and 40°C, respectively. Significant induction in the activity of intracellular oxidoreductive enzymes suggested its involvement in the decolorization of Direct Red 81. The biodegradation of Direct Red 81 was monitored by UV-Visible, FT-IR spectroscopy and HPLC. The final products were characterized by GC-MS and possible pathway of the degradation of the dye was proposed. The phytotoxicity assay (with respect to plants Sorghum vulgare and Phaseolus mungo) revealed that the degradation of Direct Red 81 produced nontoxic metabolites. Finally E. faecalis was employed to decolorize actual industrial effluent showing decolorization (in terms of ADMI value) with moderate COD and BOD reduction. Moreover the result increases the applicability of the strain for the treatment of industrial wastewaters containing dye pollutants.
Electronic supplementary material
The online version of this article (doi:10.1186/s40201-014-0151-1) contains supplementary material, which is available to authorized users.
doi:10.1186/s40201-014-0151-1
PMCID: PMC4299565  PMID: 25649265
Direct Red 81; Enterococcus faecalis; ABTS; Lignin peroxidase; Azoreductase; ADMI; GC-MS
2.  Removal of Triphenylmethane Dyes by Bacterial Consortium 
The Scientific World Journal  2012;2012:512454.
A new consortium of four bacterial isolates (Agrobacterium radiobacter; Bacillus spp.; Sphingomonas paucimobilis, and Aeromonas hydrophila)-(CM-4) was used to degrade and to decolorize triphenylmethane dyes. All bacteria were isolated from activated sludge extracted from a wastewater treatment station of a dyeing industry plant. Individual bacterial isolates exhibited a remarkable color-removal capability against crystal violet (50 mg/L) and malachite green (50 mg/L) dyes within 24 h. Interestingly, the microbial consortium CM-4 shows a high decolorizing percentage for crystal violet and malachite green, respectively, 91% and 99% within 2 h. The rate of chemical oxygen demand (COD) removal increases after 24 h, reaching 61.5% and 84.2% for crystal violet and malachite green, respectively. UV-Visible absorption spectra, FTIR analysis and the inspection of bacterial cells growth indicated that color removal by the CM-4 was due to biodegradation. Evaluation of mutagenicity by using Salmonella typhimurium test strains, TA98 and TA100 studies revealed that the degradation of crystal violet and malachite green by CM-4 did not lead to mutagenic products. Altogether, these results demonstrated the usefulness of the bacterial consortium in the treatment of the textile dyes.
doi:10.1100/2012/512454
PMCID: PMC3353484  PMID: 22623907
3.  Decolorization of the azo dye Acid Orange 51 by laccase produced in solid culture of a newly isolated Trametes trogii strain 
3 Biotech  2012;3(2):115-125.
This study concerns the decolorization and detoxification of the azo dye Acid Orange 51 (AO51) by crude laccase from Trametes trogii produced in solid culture using sawdust as support media. A three-level Box–Behnken factorial design with four factors (enzyme concentration, 1-hydroxybenzotriazole (HBT) concentration, dye concentration and reaction time) combined with response surface methodology was applied to optimize AO51 decolorization. A mathematical model was developed showing the effect of each factor and their interactions on color removal. The model predicted that Acid Orange 51 decolorization above 87.87 ± 1.27 % could be obtained when enzyme concentration, HBT concentration, dye concentration and reaction time were set at 1 U/mL, 0.75 mM, 60 mg/L and 2 days, respectively. The experimental values were in good agreement with the predicted ones and the models were highly significant, the correlation coefficient (R2) being 0.9. Then the desirability function was employed to determine the optimal decolorization condition for each dye and minimize the process cost simultaneously. In addition, germination index assay showed that laccase-treated dye was detoxified; however in the presence of HBT, the phytotoxicity of the treated dye was increased. By using cheap agro-industrial wastes, such as sawdust, a potential laccase was obtained. The low cost of laccase production may further broaden its application in textile wastewater treatment.
Electronic supplementary material
The online version of this article (doi:10.1007/s13205-012-0076-2) contains supplementary material, which is available to authorized users.
doi:10.1007/s13205-012-0076-2
PMCID: PMC3597134
Crude laccase; Synthetic textile dyes; Mediators; Optimization; Box–Behnken; Decolorization; Detoxification
4.  Decolorization and Detoxification of Textile Dyes with a Laccase from Trametes hirsuta 
Trametes hirsuta and a purified laccase from this organism were able to degrade triarylmethane, indigoid, azo, and anthraquinonic dyes. Initial decolorization velocities depended on the substituents on the phenolic rings of the dyes. Immobilization of the T. hirsuta laccase on alumina enhanced the thermal stabilities of the enzyme and its tolerance against some enzyme inhibitors, such as halides, copper chelators, and dyeing additives. The laccase lost 50% of its activity at 50 mM NaCl while the 50% inhibitory concentration (IC50) of the immobilized enzyme was 85 mM. Treatment of dyes with the immobilized laccase reduced their toxicities (based on the oxygen consumption rate of Pseudomonas putida) by up to 80% (anthraquinonic dyes). Textile effluents decolorized with T. hirsuta or the laccase were used for dyeing. Metabolites and/or enzyme protein strongly interacted with the dyeing process indicated by lower staining levels (K/S) values than obtained with a blank using water. However, when the effluents were decolorized with immobilized laccase, they could be used for dyeing and acceptable color differences (ΔE*) below 1.1 were measured for most dyes.
PMCID: PMC92155  PMID: 10919791
5.  Isolation and Characterization of Paracoccus sp. GSM2 Capable of Degrading Textile Azo Dye Reactive Violet 5 
The Scientific World Journal  2014;2014:410704.
A potential bacterial strain GSM2, capable of degrading an azo dye Reactive Violet 5 as a sole source of carbon, was isolated from textile mill effluent from Solapur, India. The 16S rDNA sequence and phenotypic characteristics indicated an isolated organism as Paracoccus sp. GSM2. This strain exhibited complete decolorization of Reactive Violet 5 (100 mg/L) within 16 h, while maximally it could decolorize 800 mg/L of dye within 38 h with 73% decolorization under static condition. For color removal, the most suitable pH and temperature were pH 6.0–9.0 and 25–40°C, respectively. The isolate was able to decolorize more than 70% of five structurally different azo dyes within 38 h. The isolate is salt tolerant as it can bring out more than 90% decolorization up to a salt concentration of 2% (w/v). UV-Visible absorption spectra before and after decolorization suggested that decolorization was due to biodegradation and was further confirmed by FT-IR spectroscopy. Overall results indicate the effectiveness of the strain GSM2 explored for the treatment of textile industry effluents containing various azo dyes. To our knowledge, this could be the first report on biodegradation of Reactive Violet 5 by Paracoccus sp. GSM2.
doi:10.1155/2014/410704
PMCID: PMC4030516  PMID: 24883397
6.  Decolorization and biodegradation of reactive sulfonated azo dyes by a newly isolated Brevibacterium sp. strain VN-15 
SpringerPlus  2012;1(1):37.
Azo dyes constitute the largest and most versatile class of synthetic dyes used in the textile, pharmaceutical, food and cosmetics industries and represent major components in wastewater from these industrial dying processes. Biological decolorization of azo dyes occurs efficiently under low oxygen to anaerobic conditions. However, this process results in the formation of toxic and carcinogenic amines that are resistant to further detoxification under low oxygen conditions. Moreover, the ability to detoxify these amines under aerobic conditions is not a wide spread metabolic activity. In this study we describe the use of Brevibacterium sp. strain VN-15, isolated from an activated sludge process of a textile company, for the sequential decolorization and detoxification of the azo dyes Reactive Yellow 107 (RY107), Reactive Black 5 (RB5), Reactive Red 198 (RR198) and Direct Blue 71 (DB71). Tyrosinase activity was observed during the biotreatment process suggesting the role of this enzyme in the decolorization and degradation process, but no-activity was observed for laccase and peroxidase. Toxicity, measured using Daphnia magna, was completely eliminated.
doi:10.1186/2193-1801-1-37
PMCID: PMC3566399  PMID: 23396675
Azo dyes; Textile wastewater; Decolorization; Biodegradation; Detoxification; Brevibacterium; Tyrosinase; Carcinogenic aromatic amine
7.  Decolorization of different textile dyes by Penicillium simplicissimum and toxicity evaluation after fungal treatment 
Brazilian Journal of Microbiology  2009;40(4):808-817.
The objective of this study was to investigate the capacity of decolorization and detoxification of the textile dyes Reactive Red 198 (RR198), Reactive Blue 214 (RB214), Reactive Blue 21 (RB21) and the mixture of the three dyes (MXD) by Penicillium simplicissimum INCQS 40211. The dye RB21, a phthalocyanine, was totally decolorized in 2 days, and the others, the monoazo RR198, the diazo RB214 and MXD were decolorized after 7 days by P. simplicissimum. Initially the dye decolorization involved dye adsorption by the biomass followed by degradation. The acute toxicity after fungal treatment was monitored with the microcrustacean Daphnia pulex and measured through Effective Concentration 50% (EC50). P. simplicissimum reduced efficiently the toxicity of RB21 from moderately acutely toxic to minor acutely toxic and it also reduced the toxicity of RB214 and MXD, which remained minor acutely toxic. Nevertheless, the fungus increased the toxicity of RR198 despite of the reduction of MXD toxicity, which included this dye. Thus, P. simplicissimum INCQS 40211 was efficient to decolorize different textile dyes and the mixture of them with a significant reduction of their toxicity. In addition this investigation also demonstrated the need of toxicological assays associated to decolorization experiments.
doi:10.1590/S1517-838220090004000011
PMCID: PMC3768592  PMID: 24031428
fungi; textile dyes; decolorization; detoxification
8.  Decolorization and detoxification of Synozol red HF-6BN azo dye, by Aspergillus niger and Nigrospora sp 
In the present investigation the fungi, Aspergillus niger and Nigrospora sp. were employed for decolorization of Synozol red HF-6BN. Decolorization study showed that Aspergillus niger and Nigrospora sp. were able to decolorize 88% and 96% Synozol red 6BN, respectively, in 24 days. It was also studied that 86% and 90% Synozol red containing of dye effluent was decolorized by Aspergillus niger and Nigrospora sp. after 28 days of incubation at room temperature. A fungal-based protein with relative molecular mass of 70 kDa was partially purified and examined for enzymatic characteristics. The enzyme exhibited highest activity at temperature ranging from 40-50°C and at pH=6.0. The enzyme activity was enhanced in the presence of metal cations. High performance liquid chromatography analysis confirmed that these fungal strains are capable to degrade Synozol red dye into metabolites. No zones of inhibition on agar plates and growth of Vigna radiata in the presence of dye extracted sample, indicated that the fungal degraded dye metabolites are nontoxic to beneficial micro-flora and plant growth. Aspergillus niger and Nigrospora sp. have promising potential in color removal from textile wastewater-containing azo dyes.
doi:10.1186/1735-2746-10-12
PMCID: PMC3605312  PMID: 23369298
Azo dyes; Decolorization; Aspergillus niger; Nigrospora sp; Bioremediation
9.  Decolorization and partial mineralization of a polyazo dye by Bacillus firmus immobilized within tubular polymeric gel 
3 Biotech  2011;2(1):67-78.
The degradation of C.I. Direct red 80, a polyazo dye, was investigated using Bacillus firmus immobilized by entrapment in tubular polymeric gel. This bacterial strain was able to completely decolorize 50 mg/L of C.I. Direct red 80 under anoxic conditions within 12 h and also degrade the reaction intermediates (aromatic amines) during the subsequent 12 h under aerobic conditions. The tubular gel harboring the immobilized cells consisted of anoxic and aerobic regions integrated in a single unit which was ideal for azo dye degradation studies. Results obtained show that effective dye decolorization (97.8%), chemical oxygen demand (COD) reduction (91.7%) and total aromatic amines removal were obtained in 15 h with the immobilized bacterial cell system whereas for the free cells, a hydraulic residence time of 24 h was required for an equivalent performance in a sequential anoxic and aerobic process. Repeated-batch experiments indicate the immobilized cells could decolorize C.I. Direct red 80 and reduce medium COD in five successive batch runs with enhanced activity obtained after each consecutive run, thus suggesting its stability and potential for repeated use in wastewater treatment. UV–visible spectrophotometry and HPLC analysis were used to confirm the partial mineralization of the dye. Data from this study could be used as a reference for the development of effective industrial scale biotechnological process for the removal of dyes and their metabolites in textile wastewater.
doi:10.1007/s13205-011-0035-3
PMCID: PMC3339580  PMID: 22582158
Decolorization; Mineralization; Azo dye; Tubular gel; Bacillus firmus; Chemistry; Bioinformatics; Agriculture; Stem Cells; Biomaterials; Biotechnology; Cancer Research
10.  Decolorization and partial mineralization of a polyazo dye by Bacillus firmus immobilized within tubular polymeric gel 
3 Biotech  2011;2(1):67-78.
The degradation of C.I. Direct red 80, a polyazo dye, was investigated using Bacillus firmus immobilized by entrapment in tubular polymeric gel. This bacterial strain was able to completely decolorize 50 mg/L of C.I. Direct red 80 under anoxic conditions within 12 h and also degrade the reaction intermediates (aromatic amines) during the subsequent 12 h under aerobic conditions. The tubular gel harboring the immobilized cells consisted of anoxic and aerobic regions integrated in a single unit which was ideal for azo dye degradation studies. Results obtained show that effective dye decolorization (97.8%), chemical oxygen demand (COD) reduction (91.7%) and total aromatic amines removal were obtained in 15 h with the immobilized bacterial cell system whereas for the free cells, a hydraulic residence time of 24 h was required for an equivalent performance in a sequential anoxic and aerobic process. Repeated-batch experiments indicate the immobilized cells could decolorize C.I. Direct red 80 and reduce medium COD in five successive batch runs with enhanced activity obtained after each consecutive run, thus suggesting its stability and potential for repeated use in wastewater treatment. UV–visible spectrophotometry and HPLC analysis were used to confirm the partial mineralization of the dye. Data from this study could be used as a reference for the development of effective industrial scale biotechnological process for the removal of dyes and their metabolites in textile wastewater.
doi:10.1007/s13205-011-0035-3
PMCID: PMC3339580  PMID: 22582158
Decolorization; Mineralization; Azo dye; Tubular gel; Bacillus firmus
11.  Decolorization of Azo, Triphenyl Methane, Heterocyclic, and Polymeric Dyes by Lignin Peroxidase Isoenzymes from Phanerochaete chrysosporium 
Applied and Environmental Microbiology  1993;59(12):4010-4016.
The ligninolytic enzyme system of Phanerochaete chrysosporium decolorizes several recalcitrant dyes. Three isolated lignin peroxidase isoenzymes (LiP 4.65, LiP 4.15, and LiP 3.85) were compared as decolorizers with the crude enzyme system from the culture medium. LiP 4.65 (H2), LiP 4.15 (H7), and LiP 3.85 (H8) were purified by chromatofocusing, and their kinetic parameters were found to be similar. Ten different types of dyes, including azo, triphenyl methane, heterocyclic, and polymeric dyes, were treated by the crude enzyme preparation. Most of the dyes lost over 75% of their color; only Congo red, Poly R-478, and Poly T-128 were decolorized less than the others, 54, 46, and 48%, respectively. Five different dyes were tested for decolorization by the three purified isoenzymes. The ability of the isoenzymes to decolorize the dyes in the presence of veratryl alcohol was generally comparable to that of the crude enzyme preparation, suggesting that lignin peroxidase plays a major role in the decolorization and that manganese peroxidase is not required to start the degradation of these dyes. In the absence of veratryl alcohol, the decolorization activity of the isoenzymes was in most cases dramatically reduced. However, LiP 3.85 was still able to decolorize 20% of methylene blue and methyl orange and as much as 60% of toluidine blue O, suggesting that at least some dyes can function as substrates for isoenzyme LiP 3.85 but not to the same extent for LiP 4.15 or LiP 4.65. Thus, the isoenzymes have different specificities towards dyes as substrates.
Images
PMCID: PMC195860  PMID: 16349103
12.  The Enzymatic Decolorization of Textile Dyes by the Immobilized Polyphenol Oxidase from Quince Leaves 
The Scientific World Journal  2014;2014:685975.
Water pollution due to release of industrial wastewater has already become a serious problem in almost every industry using dyes to color its products. In this work, polyphenol oxidase enzyme from quince (Cydonia Oblonga) leaves immobilized on calcium alginate beads was used for the successful and effective decolorization of textile industrial effluent. Polyphenol oxidase (PPO) enzyme was extracted from quince (Cydonia Oblonga) leaves and immobilized on calcium alginate beads. The kinetic properties of free and immobilized PPO were determined. Quince leaf PPO enzyme stability was increased after immobilization. The immobilized and free enzymes were employed for the decolorization of textile dyes. The dye solutions were prepared in the concentration of 100 mg/L in distilled water and incubated with free and immobilized quince (Cydonia Oblonga) leaf PPO for one hour. The percent decolorization was calculated by taking untreated dye solution. Immobilized PPO was significantly more effective in decolorizing the dyes as compared to free enzyme. Our results showed that the immobilized quince leaf PPO enzyme could be efficiently used for the removal of synthetic dyes from industrial effluents.
doi:10.1155/2014/685975
PMCID: PMC3918733  PMID: 24587743
13.  New colorimetric screening assays for the directed evolution of fungal laccases to improve the conversion of plant biomass 
BMC Biotechnology  2013;13:90.
Background
Fungal laccases are multicopper oxidases with huge applicability in different sectors. Here, we describe the development of a set of high-throughput colorimetric assays for screening laccase libraries in directed evolution studies.
Results
Firstly, we designed three colorimetric assays based on the oxidation of sinapic acid, acetosyringone and syringaldehyde with λmax of 512, 520 and 370 nm, respectively. These syringyl-type phenolic compounds are released during the degradation of lignocellulose and can act as laccase redox mediators. The oxidation of the three compounds by low and high-redox potential laccases evolved in Saccharomyces cerevisiae produced quantifiable and linear responses, with detection limits around 1 mU/mL and CV values below 16%. The phenolic substrates were also suitable for pre-screening mutant libraries on solid phase format. Intense colored-halos were developed around the yeast colonies secreting laccase. Furthermore, the oxidation of violuric acid to its iminoxyl radical (λmax of 515 nm and CV below 15%) was devised as reporter assay for laccase redox potential during the screening of mutant libraries from high-redox potential laccases. Finally, we developed three dye-decolorizing assays based on the enzymatic oxidation of Methyl Orange (470 nm), Evans Blue (605 nm) and Remazol Brilliant Blue (640 nm) giving up to 40% decolorization yields and CV values below 18%. The assays were reliable for direct measurement of laccase activity or to indirectly explore the oxidation of mediators that do not render colored products (but promote dye decolorization). Every single assay reported in this work was tested by exploring mutant libraries created by error prone PCR of fungal laccases secreted by yeast.
Conclusions
The high-throughput screening methods reported in this work could be useful for engineering laccases for different purposes. The assays based on the oxidation of syringyl-compounds might be valuable tools for tailoring laccases precisely enhanced to aid biomass conversion processes. The violuric assay might be useful to preserve the redox potential of laccase whilst evolving towards new functions. The dye-decolorizing assays are useful for engineering ad hoc laccases for detoxification of textile wastewaters, or as indirect assays to explore laccase activity on other natural mediators.
doi:10.1186/1472-6750-13-90
PMCID: PMC4015961  PMID: 24159930
High-throughput screening; Laccase; Lignocellulose; S-type phenolic mediators; Dyes; Violuric acid
14.  Optimization of culture condition for enhanced decolorization and degradation of azo dye reactive violet 1 with concomitant production of ligninolytic enzymes by Ganoderma cupreum AG-1 
3 Biotech  2012;3(2):143-152.
The strain Ganoderma cupreum AG-1 (Genbank accession no. HQ328947) isolated from the decayed wood was evaluated for its ability to decolorize azo dye reactive violet 1 as well as for the production of ligninolytic enzymes. In the initial decolorization study, the strain was capable of decolorizing 19 different azo dyes. The strain was capable of decolorizing dye over a pH range of 4.5–6 at 30 °C. The optimum pH was found to be 4.5. Various other process parameters like additional carbon and nitrogen source and initial dye concentration were also optimized. The decolorization medium was supplemented with appropriate nitrogen source (yeast extract, 5 g l−1) and carbon source (mannose, 2 g l−1); the decolorization obtained was 98 %. The pattern of enzymes involved in the biodegradation was studied and laccase and MnP were found to be the major enzymes. High laccase activity shown by G. cupreum AG-1 and its ability to decolorize dyes are a good indication of its possible use in the treatment of textile effluents.
doi:10.1007/s13205-012-0079-z
PMCID: PMC3597137
Ganoderma cupreum AG-1; Reactive violet 1; Laccase; MnP; Decolorization
15.  Enhancing the Decolorizing and Degradation Ability of Bacterial Consortium Isolated from Textile Effluent Affected Area and Its Application on Seed Germination 
The Scientific World Journal  2015;2015:628195.
A bacterial consortium BMP1/SDSC/01 consisting of six isolates was isolated from textile effected soil, sludge, and textile effluent from Hudiara drain near Nishat Mills Limited, Ferozepur Road, Lahore, Pakistan. It was selected because of being capable of degrading and detoxifying red, green, black, and yellow textile dyes. The pH and supplements were optimized to enhance the decolorization ability of the selected consortium. The results indicated that decolorizing ability of consortium for the red, green, black, and yellow dyes was higher as compared to individual strains. The consortium was able to decolorize 84%, 84%, 85%, 85%, and 82% of 200 ppm of red, green, black, yellow, and mixed dyes within 24 h while individual strain required 72 h. On supplementing urea, the consortium decolorized 87, 86, 89, 86, and 83%, respectively, while on supplementing sodium chloride the consortium decolorized 93, 94, 93, 94, and 89% of red, green, black, yellow, and mixed dyes, respectively, which was maximum while in the presence of ascorbic acid and ammonium chloride it showed intermediate results. The effect of untreated and treated dyes was investigated on Zea mays L. (maize) and Sorghum vulgare Pers. (sorghum). This study will help to promote an efficient biotreatment of textile effluents.
doi:10.1155/2015/628195
PMCID: PMC4306213  PMID: 25654132
16.  The Use of HRP in Decolorization of Reactive Dyes and Toxicological Evaluation of Their Products 
Enzyme Research  2011;2010:703824.
This work studied the potential use of horseradish peroxidase (HRP) in the decolorization of the following textile dyes: Drimarene Blue X-3LR (DMBLR), Drimarene Blue X-BLN (DMBBLN), Drimarene Rubinol X-3LR (DMR), and Drimarene Blue CL-R (RBBR). Dyes were individually tested in the reaction media containing 120 mg·L−1, considering the following parameters: temperature (20–45°C), H2O2 concentration (0–4.44 mmol·L−1), and reaction time (5 minutes, 1 and 24 h). The following conditions: 35°C, 0.55 mmol·L−1, and 1h, provided the best set of results of color removal for DMBLR (99%), DMBBLN (77%), DMR (94%), and RBBR (97%). It should be mentioned that only 5 minutes of reaction was enough to obtain 96% of decolorization for DMBLR and RBBR. After the decolorization reactions of DMBLR, DMR, and RBBR, it was possible to observe the reduction of Artemia salina mortality and the no significant increase in toxicity for the products generated from DMBBLN.
doi:10.4061/2010/703824
PMCID: PMC3034966  PMID: 21318147
17.  Decolorization of Solophenyl Red 3BL Polyazo Dye by Laccase-Mediator System: Optimization through Response Surface Methodology 
Enzyme Research  2011;2011:179050.
The decolorization of direct Solophenyl red 3BL (SR), a polyazo dye extensively used in textile industry was studied. The Fomes fomentarius laccase alone did not decolorize SR. The natural redox mediator, acetosyringone (AS), was necessary for decolorization to occur. Box-Behnken design was used to evaluate the effects of three parameters, namely, enzyme concentration (0.5–2.5 U mL−1), redox mediator concentration (3–30 μM), and incubation time (1–24 h), on the SR decolorization yield. The fitted mathematical model allowed us to plot response surfaces as well as isoresponse curves and to determine optimal decolorization conditions. The results clearly indicated that the AS concentration was the main factor influencing the SR decolorization yield. The selected optimal conditions were enzyme concentration 0.8 U mL−1, mediator concentration 33 μM, and time 14 h 30 min. These conditions allowed 79.66% of SR decolorization versus 80.70% for the predicted value. These results showed a promising future of applying laccase-AS system for industrial wastewater bioremediation.
doi:10.4061/2011/179050
PMCID: PMC3159015  PMID: 21869923
18.  Lignin-Derived Compounds as Efficient Laccase Mediators for Decolorization of Different Types of Recalcitrant Dyes 
Ten phenols were selected as natural laccase mediators after screening 44 different compounds with a recalcitrant dye (Reactive Black 5) as a substrate. Their performances were evaluated at different mediator/dye ratios and incubation times (up to 6 h) by the use of Pycnoporus cinnabarinus and Trametes villosa laccases and were compared with those of eight known synthetic mediators (including -NOH- compounds). Among the six types of dyes assayed, only Reactive Blue 38 (phthalocyanine) was resistant to laccase-mediator treatment under the conditions used. Acid Blue 74 (indigoid dye), Reactive Blue 19 (anthraquinoid dye), and Aniline Blue (triarylmethane-type dye) were partially decolorized by the laccases alone, although decolorization was much more efficient and rapid with mediators, whereas Reactive Black 5 (diazo dye) and Azure B (heterocyclic dye) could be decolorized only in the presence of mediators. The efficiency of each natural mediator depended on the type of dye to be treated but, with the only exception being Azure B (<50% decolorization), nearly complete decolorization (80 to 100%) was attained in all cases. Similar rates were attained with the best synthetic mediators, but the reactions were significantly slower. Phenolic aldehydes, ketones, acids, and esters related to the three lignin units were among the best mediators, including p-coumaric acid, vanillin, acetovanillone, methyl vanillate, and above all, syringaldehyde and acetosyringone. The last two compounds are especially promising as ecofriendly (and potentially cheap) mediators for industrial applications since they provided the highest decolorization rates in only 5 to 30 min, depending on the type of dye to be treated.
doi:10.1128/AEM.71.4.1775-1784.2005
PMCID: PMC1082544  PMID: 15812000
19.  Response surface methodology for optimization of culture conditions for dye decolorization by a fungus, Aspergillus niger HM11 isolated from dye affected soil 
Iranian Journal of Microbiology  2010;2(4):213-222.
Background and Objectives
Discharge of wastewater from textile dyeing industries has been a problem in terms of pollution and treatment of these waters is a great task. Keeping this in mind, the aim of our current research is to study the effect of various bioprocess variables on decolorization of an azo dye, Congo red, by a fungal isolate, Aspergillus niger HM11.
Materials and Methods
Central composite design (CCD) and response surface methodology (RSM) have been applied to design experiments to evaluate the interactive effects of the operating variables: on the decolorization of Congo red. A total of 30 experiments were conducted in the present study and a regression coefficient between the variables was generated.
Results
The RSM indicated that pH 6.0, 150 rpm agitation, incubation time of 36 hrs and a glucose concentration of 1.0% were optimal for maximum decolorization of Congo red and the response indicated excellent evaluation of experimental data.
Conclusion
From this study, it is very obvious that the fungal isolate, Aspergillus niger HM11 can be used as a promising microbial strain for decolorization of textile dyeing effluent containing similar dyes.
PMCID: PMC3279788  PMID: 22347575
Decolorization; Congo red; Aspergillus niger HM11; RSM; CCD
20.  Biodegradation of Green HE4B: Co-substrate effect, biotransformation enzymes and metabolite toxicity analysis 
Indian Journal of Microbiology  2010;50(2):156-164.
A high exhaust reactive dye, Green HE4B (GHE4B) was 98% degraded in nutrient medium by Pseudomonas desmolyticum NCIM 2112 (pd2112) within 72 h at static condition. Decolorization time in synthetic 10 g/l molasses. Addition of 5 g/l peptone to NaCl medium had reduced decolorization time from 108 to 72 h. Beef extract do not contribute more to the inducing effect of peptone, however it is a good co-substrate in sucrose or urea containing NaCl medium. Intracellular lignin peroxidase (Lip), laccase and tyrosinase activities were induced by 150, 355 and 212%, respectively till maximum dye removal took place. Aminopyrine N-demethylase (AND) and dichlorophenol indophenol reductase (DCIP-reductase) activities in pd2112 were induced by 130 and 20%, respectively at 72 h of incubation during GHE4B decolorization. By high performance liquid chromatography (HPLC) analysis, 4-hydroxybenzene sulfonic acid and 4-amino, 6-hydroxynaphthalene 2-sulfonic acids were identified as metabolites formed during 24–72 h incubation. Fourier transform infrared spectroscopy (FTIR) analysis supports the formation of these aromatic amines. pd2112, aerobically degraded GHE4B metabolites (formed at static condition) showing stationary phase of 6 days. There was no germination inhibition of Sorghum bicolor and Triticum aestivum by GHE4B metabolites at 3,000 ppm concentration however untreated dye showed germination inhibition at the same concentration. GHE4B metabolites did not show any microbial toxicity at 10,000 ppm concentration.
doi:10.1007/s12088-010-0001-5
PMCID: PMC3450328  PMID: 23100822
Biodegradation; Co-substrate; Green HE4B; Reactive dye; Toxicity
21.  Bacterial Decolorization of Textile Azo Dye Acid Orange by Staphylococcus hominis RMLRT03 
Toxicology International  2014;21(2):160-166.
A bacterial strain RMLRT03 with ability to decolorize textile dye Acid Orange dye was isolated from textile effluent contaminated soil of Tanda, Ambedkar Nagar, Uttar Pradesh (India). The decolorization studies were performed in Bushnell and Haas medium (BHM) amended with Acid Orange dye. The bacterial strain was identified as Staphylococcus hominis on the basis of 16S rDNA sequence. The bacterial strain exhibited good decolorization ability with glucose and yeast extract supplementation as cosubstrate in static conditions. The optimal condition for the decolorization of Acid Orange dye by Staphylococcus hominis RMLRT03 strain were at pH 7.0 and 35°C in 60 h of incubation. The bacterial strain could tolerate high concentrations of Acid Orange dye up to 600 mg l-1. The high decolorizing activity under natural environmental conditions indicates that the bacterial strain has practical application in the treatment of dye containing wastewaters.
doi:10.4103/0971-6580.139797
PMCID: PMC4170557  PMID: 25253925
16S rDNA; BHM; cosubstrate; decolorization; textile dye
22.  Decolorization of Azo Dye (Orange MR) by an Autochthonous Bacterium, Micrococcus sp. DBS 2 
Indian Journal of Microbiology  2011;51(2):159-163.
Soil and sediment samples obtained from Orange MR dye contaminated habitat were screened for heterotrophic bacterial population. The heterotrophic bacterial density of dye-contaminated soil was 2.14 × 106 CFU/g. The generic composition of heterotrophic bacterial population was primarily composed of 10% of Proteus sp., 15% Aeromonas sp., 20% Bacillus sp., 25% Pseudomonas sp. and 30% Micrococcus sp. The bacterial strain that decolorized the azo dye Orange MR up to 900 ppm was identified as Micrococcus sp. The optimum inoculum load, pH and temperature were found to be 5%, 6 and 35°C, respectively. The rate of decolorization was assessed using spectrophotometer at 530 nm and the percentage of decolorization was ascertained. The autochthonous bacterial isolate was able to utilize the dye as both nitrogen and carbon source.
doi:10.1007/s12088-011-0127-0
PMCID: PMC3209883  PMID: 22654158
Azo dye; Orange MR; Autochthonous bacteria; Micrococcus; Decolorization
23.  Recent Advances in Heterogeneous Photocatalytic Decolorization of Synthetic Dyes 
The Scientific World Journal  2014;2014:692307.
During the process and operation of the dyes, the wastes produced were commonly found to contain organic and inorganic impurities leading to risks in the ecosystem and biodiversity with the resultant impact on the environment. Improper effluent disposal in aqueous ecosystems leads to reduction of sunlight penetration which in turn diminishes photosynthetic activity, resulting in acute toxic effects on the aquatic flora/fauna and dissolved oxygen concentration. Recently, photodegradation of various synthetic dyes has been studied in terms of their absorbance and the reduction of oxygen content by changes in the concentration of the dye. The advantages that make photocatalytic techniques superior to traditional methods are the ability to remove contaminates in the range of ppb, no generation of polycyclic compounds, higher speed, and lower cost. Semiconductor metal oxides, typically TiO2, ZnO, SnO, NiO, Cu2O, Fe3O4, and also CdS have been utilized as photocatalyst for their nontoxic nature, high photosensitivity, wide band gap and high stability. Various process parameters like photocatalyst dose, pH and initial dye concentrations have been varied and highlighted. Research focused on surface modification of semiconductors and mixed oxide semiconductors by doping them with noble metals (Pt, Pd, Au, and Ag) and organic matter (C, N, Cl, and F) showed enhanced dye degradation compared to corresponding native semiconductors. This paper reviews recent advances in heterogeneous photocatalytic decolorization for the removal of synthetic dyes from water and wastewater. Thus, the main core highlighted in this paper is the critical selection of semiconductors for photocatalysis based on the chemical, physical, and selective nature of the poisoning dyes.
doi:10.1155/2014/692307
PMCID: PMC4099358  PMID: 25054183
24.  Decolorization applicability of sol–gel matrix immobilized manganese peroxidase produced from an indigenous white rot fungal strain Ganoderma lucidum 
BMC Biotechnology  2013;13:56.
Background
An eco-friendly treatment of industrial effluents is a major environmental concern of the modern world in the face of stringent environmental legislations. By keeping in mind the extensive industrial applications of ligninolytic enzymes, this study was performed to purify, and immobilize the manganese peroxidase (MnP) produced from an indigenous strain of Ganoderma lucidum. The present study was also focused on investigating the capability of immobilized MnP for decolorization of dye containing textile effluents.
Results
A large magnitude of an indigenous MnP (882±13.3 U/mL) was obtained from white rot fungal strain G. lucidum in solid state bio-processing of wheat straw under optimized fermentation conditions (moisture, 50%; substrate, 5 g; pH, 5.5; temperature, 30°C; carbon source, 2% fructose; nitrogen source, 0.02% yeast extract; C: N ratio, 25:1; fungal spore suspension, 5 mL and fermentation time period, 4 days). After ammonium sulfate fractionation and Sephadex-G-100 gel filtration chromatography, MnP was 4.7-fold purified with specific activity of 892.9 U/mg. G. lucidum MnP was monomeric protein as evident by single band corresponding to 48 kDa on native and denaturing SDS-PAGE. The purified MnP (2 mg/mL) was immobilized using a sol–gel matrix of tetramethoxysilane (TMOS) and proplytrimethoxysilane (PTMS). The oxidation of MnSO4 for up to 10 uninterrupted cycles demonstrated the stability and reusability of the immobilized MnP. Shelf life profile revealed that enzyme may be stored for up to 60 days at 25°C without losing much of its activity. To explore the industrial applicability of MnP produced by G. lucidum, the immobilized MnP was tested against different textile effluents. After 4 h reaction time, the industrial effluents were decolorized to different extents (with a maximum of 99.2%). The maximally decolorized effluent was analyzed for formaldehyde and nitroamines and results showed that the toxicity parameters were below the permissible limits.
Conclusions
In conclusion, G. lucidum MnP was immobilized by sol–gel matrix entrapment with an objective to enhance its practical efficiencies. The MnP was successfully entrapped into a sol- gel matrix of TMOS and PTMS with an overall immobilization efficiency of 93.7%. The sol- gel entrapped MnP seems to have prospective capabilities which can be useful for industrial purposes, especially for bioremediation of industrial effluents.
doi:10.1186/1472-6750-13-56
PMCID: PMC3717284  PMID: 23849469
Bio-catalysis; G. lucidum; MnP; PAGE; Sol–gel; Immobilization; Textile effluents; Decolorization; Toxicity reduction
25.  A New Alkali-Thermostable Azoreductase from Bacillus sp. Strain SF 
A screening for dye-decolorizing alkali-thermophilic microorganisms resulted in a Bacillus sp. strain isolated out of the wastewater drain of a textile finishing company. An NADH-dependent azoreductase of this strain, Bacillus sp. strain SF, was found to be responsible for the decolorization of azo dyes. This enzyme was purified by a combination of ammonium sulfate precipitation and anion-exchange and affinity chromatography and had a molecular mass of 61.6 kDa and an isoelectric point at pH 5.3. The pH optimum of the azoreductase depended on the substrate and was within the range of pHs 8 to 9, while the temperature maximum was reached at 80°C. Decolorization only took place in the absence of oxygen and was enhanced by FAD, which was not consumed during the reaction. A 26% similarity of this azoreductase to chaperonin Cpn60 from a Bacillus sp. was found by peptide mass mapping experiments. Substrate specificities of the azoreductase were studied by using synthesized model substrates based on di-sodium-(R)-benzyl-azo-2,7-dihydroxy-3,6-disulfonyl-naphthaline. Those dyes with NO2 substituents, especially in the ortho position, were degraded fastest, while analogues with a methyl substitution showed the lowest degradation rates.
doi:10.1128/AEM.70.2.837-844.2004
PMCID: PMC376287  PMID: 14766562

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