Laccase-membrane reactors for decolorization of an acid azo dye in aqueous phase: Process optimization

In the present investigation, performance of various laccase-membrane reactor configurations including direct enzyme contact, enzyme impregnated, immobilized enzyme and a reactor system based on laccase immobilization in chitosan membranes for decolorization of azo dye (acid black 10 BX) were examined using laccase enzyme purified from white rot fungi Pleurotus ostreatus 1804. A five-step laccase purification procedure was employed, which improved the enzymatic activity by 8.27 folds. Laccase was confirmed by comparing with the standard marker using SDS-PAGE electrophoresis, which showed molecular weight of 63 kDa. Experimental data showed that laccase has great potential for color removal without addition of external redox mediators. Various process parameters viz. aqueous phase of pH 6.0, enzyme concentration of 1.75 U/ml, dye concentration of 20 mg/L, temperature of 30 °C and reaction time of 120 min were optimized to achieve maximum decolorization efficiencies. Moreover, different laccase-membrane reactor configurations were tested to determine the efficacy of repeated application of laccase on dye decolorization process. Among the different reactor configurations employed, laccase encapsulated in chitosan membrane showed advantages such as short-term contact period and reusability of enzyme for a number of cycles.     

Decolorization of textile dyes by laccases from a newly isolated strain of Trametes modesta

Four ligninolytic fungi, Trametes modesta, Trametes hirsuta, Trametes versicolor and Sclerotium rolfsii, were compared for their ability to produce laccases. The fungal laccases were screened for their ability to decolorize eight synthetic dyes (anthraquinone, azo, indigo and triarylmethane). The decolorization rate depended both on the source of the enzyme preparation and on the structure of the dye. Based on laccase production and dye decolorizing ability, T. modesta was selected for further studies. All the tested dyes were decolorized by the T. modesta laccase most efficiently under acid conditions (pH 3-6) but the optimum pH for decolorization of the individual dye varied. The decolorization rate of this laccase increased with the rise in temperature to 50-60 degrees C. The decolorization efficiency of T. modesta laccase was improved remarkably in the presence of mediators like 1-hydroxybenzotriazole and 2-methoxyphenothiazine.     

Degradation of a textile reactive Azo dye by a combined chemical-biological process: Fenton's reagent-yeast

This work presents the results of our studies on the decolorization of aqueous azo dye Reactive black 5 (RB5) solution combining an advanced oxidation process (Fenton's reagent) followed by an aerobic biological process (mediated by the yeast Candida oleophila). Under our conditions, initial experiments showed that Fenton's process alone, as well as aerobic treatment by C. oleophila alone, exhibited the capacity to significantly decolorize azo dye solutions up to 200 mg/L, within about 1 and 24h, respectively. By contrast, neither Fenton's reagent nor C. oleophila sole treatments showed acceptable decolorizing abilities for higher initial dye concentrations (300 and 500 mg/L). However, it was verified that Fenton's reagent process lowered these higher azo dye concentrations to a value less than 230 mg/L, which is apparently compatible with the yeast action. Therefore, to decolorize higher concentrations of RB5 and to reduce process costs the combination between the two processes was evaluated. The final decolorization obtained with Fenton's reagent process as primary treatment, at 1.0 x 10(-3)mol/L H(2)O(2) and 1.0 x 10(-4)mol/L Fe(2+), and growing yeast cells as a secondary treatment, achieves a color removal of about 91% for an initial RB5 concentration of 500 mg/L.     

Mechanism of textile metal dye biotransformation by Trametes versicolor

The biodegradation of Grey Lanaset G, which consists of a mixture of metal complexed dye, was studied. Experiments were carried out in a bioreactor with retained pellets of the fungus Trametes versicolor that was operated under conditions of laccase production. Although decolorization was highly efficient (90%), no direct relationship to extracellular enzyme was apparent. Moreover, the extracellular enzyme was found to be unable to degrade the dye in vitro. The process involves several steps. Thus, the initial adsorption of the dye and its transfer into cells is followed by breaking of the metal complex bond in the cells release of the components. The metal (Cr and Co) contents of the biomass and treated solutions, and their closer relationship to intracellular enzyme and degradation of the dye, confirm the initial hypothesis.     

Decolorization of an azo dye by unacclimated activated sludge under anaerobic conditions

Studies on the reductive decolorization of a complex azo dye, Reactive Red 3.1, were made as part of the development of a practical approach to better exploit the metabolic potential of biomass in wastewater treatment. Decolorization was achieved at low and variable rates by mixed microbial cultures under various environmental conditions, including low pH and high salt concentration. It was caused by reductive cleavage of the azo bond to yield two aromatic amines. More reliable and effective decolorization rates, of up to 20–30 mg l⁻¹ h⁻¹, were given by unadapted activated sludge, (6 g l⁻¹) incubated with 400 mg l⁻¹ of Reactive Red 3.1 under anaerobic conditions. Decolorization also occurred best in static conditions.     

The photocatalytic degradation of reactive black 5 using TiO2/UV in an annular photoreactor

The textile effluent is a major industrial polluter because it is highly colored, containing about 15% unfixed dyes as well as high levels of salts that can potentially be discharged into the environment. Photocatalytic oxidation using an thin gap annular UV reactor with TiO2 was used to break down the colour of a synthetic effluent ranging up to 400 ppm in dye concentration of Reactive Black 5 and up to 80 g/L in NaCl. Results show that the reaction kinetics was dominated by the TiO2 loading, the initial dye concentration, and the dissolved oxygen concentration; with the other parameters showing less significant effects. High rates of decolorization were found, with a linear fit to the Langmuir-Hinshelwood equation yielding a reaction rate constant (k) of 2.45 ppm/min, and an adsorption equilibrium constant (K) of 0.048 ppm(-1) based on color removal. The presence of the combination of high dissolved oxygen (15 ppm) and sodium chloride (up to 80 g/L) was found to enhance the decolorization and mineralization rates of the reactive dye. However, pH was found to not significantly affect the degradation rate. Since textile effluent is strongly alkaline, this result is significant, as no solution neutralisation is required and direct treatment of the effluent is possible.     

Enzymatic biodegradation and detoxification of azo and anthraquinone dyes by indigenously isolated bacterial monocultures and their synergistic behaviour in developed consortia

Wastewater generated by textile industry needs to be treated to reduce its toxicity before final disposal and/or for recycling purposes. In the current study, several bacterial strains were screened for dye decolorization potential. UV–visible spectroscopy was used to determine maximum absorption wavelength of disperse dyes. HPLC and MTS assay were used to confirm the degradation and detoxification of disperse dyes, respectively. Results revealed that indigenously isolated Bacillus licheniformis, Glutamicibacter uratoxydans and Pseudomonas aeruginosa showed strong decolorization of red, blue and violet, respectively in 6–9 h. MTS assay revealed 100% viability of NIH/3T3 cell lines in presence of treated dyes. Enzyme screening assay confirmed the production of intracellular and membrane bound oxidoreductases in presence of specific dye as substrate. To resolve this issue, bacterial consortia were prepared, and better decolorization of all dyes was achieved in synergistic behaviour of Consortia 1 and 4 with 85% and 88% decolorization potential, respectively.     

Comparative studies on potential of consortium and constituent pure bacterial isolates to decolorize azo dyes

The decolorization potential of the consortium HM-4 constituted by mixing four laboratory isolates identified as Bacillus cereus (BN-7), Pseudomonas putida (BN-4), Pseudomonas fluorescens (BN-5) and Stenotrophomonas acidaminiphila (BN-3) was compared with that of individual isolates. Six different azo dyes viz., C.I. Acid Red 88 (AR-88), C.I. Acid Red 119 (AR-119), C.I. Acid Red 97 (AR-97), C.I. Reactive Red 120 (RR-120), C.I. Acid Blue 113 (AB-113) and C.I. Acid Brown 100 (AB-100) were used in this study. The individual bacterial isolates were not able to completely decolorize these dyes, except for dyes AR-119 and AB-113. The consortium HM-4 was able to decolorize all the dyes used at an initial dye concentration of 20 mg L⁻¹ at a significantly higher rate as compared to that achieved by individual isolates.     

Further treatment of decolorization liquid of azo dye coupled with increased power production using microbial fuel cell equipped with an aerobic biocathode

A microbial fuel cell (MFC) incorporating a recently developed aerobic biocathode is designed and demonstrated. The aerobic biocathode MFC is able to further treat the liquid containing decolorization products of active brilliant red X-3B (ABRX3), a respective azo dye, and also provides increased power production. Batch test results showed that 24.8% of COD was removed from the decolorization liquid of ABRX3 (DL) by the biocathode within 12 h. Metabolism-dependent biodegradation of aniline-like compound might be mainly responsible for the decrease of overall COD. Glucose is not necessary in this process and contributes little to the COD removal of the DL. The similar COD removal rate observed under closed circuit condition (500 Ω) and opened circuit condition indicated that the current had an insignificant effect on the degradation of the DL. Addition of the DL to the biocathode resulted in an almost 150% increase in open cycle potential (OCP) of the cathode accompanied by a 73% increase in stable voltage output from 0.33 V to 0.57 V and a 300% increase in maximum power density from 50.74 mW/m² to 213.93 mW/m². Cyclic voltammetry indicated that the decolorization products of the ABRX3 contained in the DL play a role as redox mediator for facilitating electron transfer from the cathode to the oxygen. This study demonstrated for the first time that MFC equipped with an aerobic biocathode can be successfully applied to further treatment of effluent from an anaerobic system used to decolorize azo dye, providing both cost savings and high power output.     

Biodegradation and detoxification potential of rotating biological contactor (RBC) with Irpex lacteus for remediation of dye-containing wastewater

Use of fungal organisms in rotating biological contactors (RBC) for bioremediation of liquid industrial wastes has so far been limited in spite of their significant biodegradation potential. The purpose was to investigate the power of RBC using Irpex lacteus for decolorization and detoxification of industrial dyes and dyeing textile liquors. Recalcitrant dye Methylene Blue (150 mg L⁻¹) was decolorized within 70 days, its mutagenicity removed, and the biological toxicity decreased more than 10-fold. I. lacteus biofilm in the RBC completely decolorized within 26 and 47 days dyeing liquors containing disperse or reactive dyes adjusted to pH4.5 and 5-fold diluted with the growth medium, respectively. Their respective biological toxicity values were reduced 10- to 10⁴-fold in dependence of the test used. A battery of toxicity tests comprising Vibrio fisheri, Lemna minor and Sinapis alba was efficient to monitor the toxicity of textile dyes and wastewaters. Strong decolorization and detoxification power of RBC using I. lacteus biofilms was demonstrated.     

Decolorization and toxicity of reactive anthraquinone textile dyes under methanogenic conditions

Reductive decolorization of two anthraquinone reactive dyes (Reactive Blue 4, RB4; Reactive Blue 19, RB19) under methanogenic conditions was performed using a mixed, methanogenic culture. Decolorization of the two anthraquinone dyes was investigated to evaluate the rate and extent of color removal as well as to assess possible toxic effects of the dyes and their decolorization product(s) on the methanogenic culture as a function of initial dye concentration ranging from 50 to 300 mg x L(-1). A dextrin/peptone mixture was used as the carbon and electron source. A high rate and extent of color removal was achieved ranging from 4.3 to 29.9 mg x L(-1)h(-1) and 73-91% for RB4, and 13.0-74.4 mg x L(-1)h(-1) and 90-95% for RB19. Initial RB4 concentrations up to 100 mg x L(-1) did not result in any significant inhibition. Both the 200 and 300 mg x L(-1) RB4-amended cultures, and all RB19-amended cultures resulted in severe inhibition of both acidogenesis and methanogenesis. Sequential dye addition at 300 mg x L(-1) for both RB4 and RB19 resulted in accumulation of volatile fatty acids (VFAs) and a very low methane production at the end of the first dye addition after 44 days of incubation. However, at the end of the second dye addition, after a relatively long incubation (384 days), recovery of methanogens in the RB4-amended culture was observed in contrast to the complete inhibition of methanogenesis in the RB19-amended culture. Therefore, RB19 resulted in a higher degree of inhibition of both acidogenesis and methanogenesis than RB4. Addition of dextrin/peptone to dye-inhibited cultures resulted in acidogenesis and a gradual recovery of methanogenesis (mainly aceticlastic methanogenesis) in the RB4-inhibited culture, and a slow recovery of acidogenesis but no recovery of methanogenesis in the RB19-inhibited culture. In contrast, addition of 80% H(2)-20% CO(2) gas to dye-inhibited cultures resulted in recovery of hydrogenotrophic methanogenesis in both the RB4- and RB19-inhibited cultures. In spite of the relatively severe inhibition of the two anthraquinone dyes on the mixed, methanogenic culture, a high extent of color removal was achieved.     

Preparation and photocatalytic performance of Fe (III)-amidoximated PAN fiber complex for oxidative degradation of azo dye under visible light irradiation

Polyacrylonitrile (PAN) fiber was modified with hydroxylamine hydrochloride to introduce amidoxime groups onto the fiber surface. These amidoxime groups were used to react with Fe (III) ions to prepare Fe (III)-amidoximated PAN fiber complex, which was characterized using SEM, XRD, FTIR, XPS, DMA, and DRS respectively. Then the photocatalytic activity of Fe-AO-PAN was evaluated in the degradation of a typical azo dye, C. I. Reactive Red 195 in the presence of H₂O₂ under visible light irradiation. Moreover, the effect of the Fe content of Fe-AO-PAN on dye degradation was also investigated. The results indicated that Fe (III) ions can crosslink between the modified PAN fiber chains by the coordination of Fe (III) ions with the amino nitrogen atoms and hydroxyl oxygen atoms of the amidoximation groups to form Fe (III)-amidoximated PAN fiber complex, and the Fe content of which is mainly determined by Fe (III) ions and amidoximation groups. Fe (III)-amidoximated PAN fiber complex is found to be activated in the visible light region. Moreover, Fe (III)-amidoximated PAN fiber complex shows the catalytic activity for dye degradation by H₂O₂ at pH = 6.0 in the dark, and can be significantly enhanced by increasing light irradiation and Fe content, therefore, it can be used as a new heterogeneous Fenton photocatalyst for the effective decomposition of the dye in water. In addition, ESR spectra confirm that Fe (III)-amidoximated PAN fiber complex can generate more OH radicals from H₂O₂ under visible light irradiation, leading to dye degradation. A possible mechanism of photocatalysis is proposed.     

Kinetic characteristics of bacterial azo-dye decolorization by Pseudomonas luteola

A Pseudomonas luteola strain expressing azoreductase activity was utilized to remove the color of an azo dye (reactive red 22) from contaminated solutions. The effects of substrate concentrations, medium compositions, and operation parameters (e.g., pH, temperature, dissolved oxygen, etc.) on decolorization of the azo dye by a P. luteola strain were systematically investigated to reveal the key factors that dominate the performance of azo-dye decolorization. The metabolites resulting from bacterial decolorization were analyzed by high-performance liquid chromatography (HPLC) and mass spectrometery (MS). The results show that the dissolved oxygen and glucose concentration retarded decolorization of reactive red 22 by P. luteola. The optimal azo-dye decolorization occurred at 37 degrees C, while more rapid decolorization took place over pH 7-9. Yeast extract and tryptone strongly enhanced the decolorization. The Michaelis-Menten model can satisfactorily describe the dependence of specific decolorization rate on the concentration of substrate (reactive red 22 or yeast extract). Decolorization of the azo dye by intact cells of P. luteola was essentially independent of the growth phase, whereas the azoreductase activity of the cell-free extract decreased in the order of late-stationary phase > early-stationary phase > mid-log phase. This suggests that mass transfer of the azo dye across the cell membrane may be the rate-limiting step. The HPLC and MS analyses suggest that both partial reduction and complete cleavage of the azo bond could contribute to decolorization of reactive red 22 by P. luteola.     

Biodegradation potential of pure and mixed bacterial cultures for removal of 4-nitroaniline from textile dye wastewater

Environmentally toxic aromatic amines including nitroanilines are commonly generated in dye contaminated wastewater in which azo dyes undergo degradation under anaerobic conditions. The aim of this study was to develop a process for biological treatment of 4-nitroaniline. Three bacteria identified as Acinetobacter sp., Citrobacter freundii and Klebsiella oxytoca were isolated from enrichment cultures of activated sludge on 4-nitroaniline, after which the isolates and the mixed culture were studied to determine optimal conditions for biodegradation. HPLC analyses showed the mixed culture was capable of complete removal of 100 μmol/L of 4-nitroaniline within 72 h under aerobic conditions. There was an inverse linear relationship (R² = 0.96) between the rate of degradation (V) and 4-nitraoaniline concentrations [S] over 100-1000 μmol/L. The bacterial culture was also capable of decolorizing structurally different azo dyes (Acid Red-88, Reactive Black-5, Direct Red-81, and Disperse Orange-3) and also degraded nitrobenzene. Our findings show that enrichment cultures from activated sludge can be effective for the removal of dyes and their toxic intermediates, and that treatment may best be accomplished using an anaerobic-aerobic process.     

Investigation of isolation and immobilization of a microbial consortium for decoloring of azo dye 4BS

Eight high-effective decolorization strains were isolated by enrichment using Direct Fast Scarlet 4BS as sole source of carbon and energy. The optimal microbial consortium consisting of fungus 8-4(*) and bacterium 1-10 was selected by optimizing combination decolorization experiments with these eight freely suspended strains, whose decolorization activity was higher than individual strains due to synergistic reaction with each other. The optimal microbial consortium was also immobilized using polyvinyl alcohol (PVA) as the carrier. This paper optimized the immobilization and operational conditions, investigated the effect of the environmental factors (temperature, pH and dissolved oxygen (DO)) and initial dye concentration on the rate of decolorization by immobilized microbial consortium. The results showed that the optimal decolorization activity was observed in pH range (5-8), temperature range (25-40 degrees C) under shaking culture of high DO level. Decolorization with the optimal microbial consortium gave a relatively high maximum decolorization activity (ca. 81.25 mgl(-1)h(-1)), which occurred at a dye concentration of 1000 mgl(-1), suggesting the applicability of the strains in remediation of wastewater containing high azo dye concentrations. The immobilized beads could be reused for more than 30 cycles, without losing any degradation capacity. The changes of UV-visible spectra and the change curve of COD of 4BS solution before and after decolorization cultivation and the proliferation and distribution of microbial consortium in gel beads were also microscopically observed, which could be used for conferring the decolorization mechanisms of dye 4BS.     

Optimal decolorization and kinetic modeling of synthetic dyes by Pseudomonas strains

Pseudomonas spp were isolated from an anaerobic-aerobic dyeing house wastewater treatment facility as the most active azo-dye degraders. Decolorization of azo dyes and non-azo dyes including anthraquinone, metal complex and indigo was compared with individual strains and a bacterial consortium consisting of the individual strain and municipal sludge (50 50wt). The consortium showed a significant improvement on decolorization of two recalcitrant non-azo dyes, but little effect on the dyes that the individual strains could degrade to a great or moderate extent. Decolorization of Acid violet 7 (monoazo) by a Pseudomonas strain GM3 was studied in detail under various conditions. The optimum decolorization activity was observed in a narrow pH range (7-8), a narrow temperature range (35-40 degrees C), and at the presence of organic and ammonium nitrogen. Nitrate had a severe inhibitory effect on azo dye decolorization: 10 mg/L led to 50% drop in decolorization activity and 1000 mg/L to complete activity depression. A kinetic model is established giving the dependence of decolorization rate on cell mass concentration (first-order) and dye concentration (half order). The rate increased with temperature from 10 to 35 C, which can be predicted by Arrhenius equation with the activation energy of 16.87 kcal/mol and the frequency factor of 1.49 x 10(11) (mg L)1/2/g DCM min.     

Application of 'waste' metal hydroxide sludge for adsorption of azo reactive dyes

The capacity and mechanism of metal hydroxide sludge in removing azo reactive dyes from aqueous solution was investigated with different parameters, such as charge amount of dyes, system pH, adsorbent particle size, and adsorbent dosage. The three anionic dyes used were CI Reactive Red 2, CI Reactive Red 120, and CI Reactive Red 141, increasing in number of sulfonic groups, respectively. Only 0.2% (w/v) of powdered sludge (<75microm) achieved color removal from 30 mg l(-1) reactive dye solutions within 5 min without pH adjustment. The larger the charge amount of the dyes, the greater the adsorption (>90%) on the metal hydroxide sludge. The system pH played a significant role in the adsorption on metal hydroxides and formation of dye-metal complexes. The optimum system pH for dye adsorption was 8-9 which was close to the pH(zpc) of the sludge while the precipitation of dye-metal complexes occurred at system pH 2. The maximum adsorption capacity (Q degrees ) of the sludge for the reactive dyes was 48-62 mg dye g(-1) adsorbent. The Langmuir and Freundlich models showed that the higher charged dyes had a higher affinity of adsorption. The smaller particle size and the greater amount of adsorbent showed the faster process, due to an increase in surface area of adsorbent. Desorption studies elucidated that metal hydroxide sludge had a tendency for ion exchange adsorption of sulfonated azo reactive dyes. Leaching data showed that the treated water was nontoxic at a system pH above 5 or a solution pH above 2.     

Kinetic modeling and synergy quantification in sono and photooxidative treatment of simulated dyehouse effluent

The aim of this work was to explore the application of sulfate radical based advanced oxidation processes: photooxidation (UV/PMS/PS), sonooxidation (US/PMS/PS) and combined sono-photooxidation (US/UV/PMS/PS) for the mineralization of simulated dyehouse effluent (WW); using peroxymonosulfate (PMS) and persulfate (PS) as oxidants. Experiments were performed in a reaction vessel of a defined geometry and axially positioned source of UV-C radiation, all placed in the ultrasonic bath (35 kHz). Mathematical model of the process was developed according to the proposed degradation scheme. Decomposition of dyestuff (C.I. Reactive Violet 2, RV2 and C.I. Reactive Blue 7, RB7), surfactant (linear alkylbenzene sulfonate; hereafter: LAS) and auxiliary organic components was explored in three types of model wastewater: WW, simulated effluent excluding inorganic species (WW-IS) and model solution that consists of a specific compound (hereafter: compound model solutions). The influence of inorganic matrix (Cl⁻, CO₃²⁻/HCO₃⁻) was studied due to the corresponding quenching affinity toward HO and SO₄⁻ radicals. The efficiency of applied processes was evaluated and the response to combined phenomena (cavitation and irradiation) was quantified as synergy index, f_Syn. Sono-photooxidative treatment (US/UV/PMS/PS) of WW resulted in a partial mineralization and partial decolourization; approximately 40% of initial TOC and 30% of initial RB7 remained after 60 min of treatment, while RV2 and LAS molecule were completely decomposed. Circumstantially, the combined process increased the mineralization efficiency by a factor of 3 (f_Syn = 3.026).     

Bioremediation of crystal violet using air bubble bioreactor packed with Pseudomonas aeruginosa

Seven water and sediment samples were collected and tested for decolorizing crystal violet. Pseudomonas aeruginosa was the most effective isolate for dye decolorization. The LC₅₀ of the crystal violet (115 mg/l) was measured using Artemia salina as a biomarker. The effect of different heavy metals on crystal violet decolorization was investigated. Cd²⁺ and Fe³⁺ ions showed marginal enhancement of the decolorization process, the rate was 1.35 mg/l/h compared to (1.25 mg/l/h) for the control. Phenol and m-cresol showed no effect on crystal violet decolorization, meanwhile p-cresol and p-nitrophenol reduced the decolorization rate to 1.07 and 0.01 mg/l/h, respectively. P. aeruginosa cells were immobilized by entrapment in agar-alginate beads. The beads were cultivated and reused in Erlenmeyer flask and in an air bubble column bioreactor and they enhanced the crystal violet decolorization rate to 3.33 and 7.5 mg/l/h, respectively.     

Bioremediation of crystal violet using air bubble bioreactor packed with Pseudomonas aeruginosa

Seven water and sediment samples were collected and tested for decolorizing crystal violet. Pseudomonas aeruginosa was the most effective isolate for dye decolorization. The LC₅₀ of the crystal violet (115 mg/l) was measured using Artemia salina as a biomarker. The effect of different heavy metals on crystal violet decolorization was investigated. Cd²⁺ and Fe³⁺ ions showed marginal enhancement of the decolorization process, the rate was 1.35 mg/l/h compared to 1.25 mg/l/h for the control. Phenol and m-cresol showed no effect on crystal violet decolorization, meanwhile p-cresol and p-nitrophenol reduced the decolorization rate to 1.07 and 0.01 mg/l/h, respectively. P. aeruginosa cells were immobilized by entrapment in agar–alginate beads. The beads were cultivated and reused in Erlenmeyer flask and in an air bubble column bioreactor and they enhanced the crystal violet decolorization rate to 3.33 and 7.5 mg/l/h, respectively.