batch reactor. N.D. Lourenço, J.M. Novais and H.M. Pinheiro. Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 ...
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N.D. Lourenço, J.M. Novais and H.M. Pinheiro Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal Abstract Two sequencing batch reactors (SBRs) with sequenced anaerobic/aerobic phases were used to study biological colour removal from a simulated cotton textile effluent containing an azo reactive dye. One of the reactors was daily fed with Remazol Brilliant Violet 5R dye and the other was used as control. When operating with a sludge retention time (SRT) of 15 days the total COD removal was around 80%, with 30% being removed anaerobically. After 40–50 days of acclimatization the colour removal efficiency reached a maximum, stable value of 90% from a feed dye concentration of 90 mg/l, almost all being removed during the anaerobic phase. This colour removal was attributed to microbial degradation rather than adsorption and colour removal capacity was not lost even after a seven-day absence of dye in the fed substrate. The dye-fed reactor experienced a reduction in the ORP values attained during the non-aerated phase, after acclimatization, an effect not observed in the dye-free control. Under denitrifying conditions it was observed that the decolouration levels achieved in the anaerobic phase decreased from 90% to 70% after only two cycles with a feed containing 45–60 mg NO3/l. Reduction of the SRT value from 15 to 10 days reduced the biomass concentration from 2.0 to 1.2 g VSS/l and lowered colour removal levels from 90% to 30–50%. When the SRT value was increased back to 15 days the colour removal capacity of the system was completely recovered, suggesting that with a SRT of 10 days the adequate microbial population could not be installed in the reactors. Keywords Anaerobic-aerobic treatment; azo dyes; colour removal; reactive dyes; SBR; textile effluents
Water Science and Technology Vol 42 Nos 5–6 pp 321–328 © IWA Publishing 2000
Reactive textile dye colour removal in a sequencing batch reactor
Introduction
Reactive dyes have widespread use in the food, cosmetic, paper and cotton textile industries and among them, the most common used chromophores are azo-based, owing to their structural versatility, ability to bind to most synthetic and natural fibers and their potential to cover the entire visible spectrum (Bumpus, 1995). Heavy colour can result in the wastewaters of textile manufacturing industries using reactive azo dyes owing to their high water solubility and to the significant fraction of the dye that is hydrolysed during the dyeing process and does not bind to the fibers (Ganesh et al., 1994). Color removal from textile effluents is still a major environmental concern because reactive azo dyes are not readily degradable and are consequently difficult to remove by the conventional wastewater treatment systems (Pagga and Brown, 1986; Seshadri et al., 1994; Bumpus, 1995). There are several physical and chemical techniques available for the treatment of coloured effluents, including adsorption by activated carbon, coagulation and sedimentation, bleaching with chlorine or ozone and reverse osmosis (Knapp et al., 1997), but their cost is the major drawback of these techniques (Churchley, 1994). On the other hand a biological treatment system normally presents lower operating costs and improved applicability (Beydilli et al., 1998). Many of the reported examples of azo dye decolouration by microorganisms take place under anaerobic conditions starting with the formation of intermediary aromatic amines by reductive cleavage of the azo bond (Walker, 1970; Wuhrmann et al., 1980; Haug et al., 1991; Voyksner et al., 1993; Kudlich et al., 1996). Though this step removes the colour of the dye it does not remove the dye-related hazard from the wastewater since anaerobic
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mineralization of most of the resulting aromatic amines has not been reported, with the exception of a few hydroxyl and carboxyl substituted amines that were completely degraded under methanogenic conditions (Razo-Flores et al., 1996; 1997). Some of these amines can be toxic or carcinogenic (Pasti-Grigsby et al., 1992; Field et al., 1995; Carliell et al., 1998). However, it has been reported that many of them can be degraded under aerobic conditions by a hydroxylation and ring opening mechanism (Bumpus, 1995; Zissi and Lyberatos, 1996). Recently a combination of anaerobic and aerobic steps has been used in the attempt to achieve both decolouration and mineralization of azo reactive dyes (Haug et al., 1991; Zaoyan et al., 1992; Seshadri et al., 1994; Kudlich et al., 1996; Hu, 1998; Krull et al., 1998). In the present work a sequencing batch reactor operating with sequenced anaerobic-aerobic steps was chosen to study colour removal from a simulated cotton textile effluent containing a reactive azo dye. Materials and method Hydrolysed Emsize E1 preparation
Emsize E1 (Emsland-Stärke GmbH, Germany) is a starch derivative used as a sizing agent in the cotton textile industry. Under the alkaline conditions applied in the desizing step of cotton manufacture the sizing agent is hydrolysed and released in the effluent. The preparation of the hydrolysed solution (100 g/l) was based on one set of desizing conditions provided by the manufacturer. 100 g of Emsize E1 and 40 g of sodium hydroxide were dissolved in distilled water and stirred during 15 hours at room temperature. The hydrolysed solution was then neutralized to pH 7.0±0.05 with 37% HCl and diluted to 1 litre with distilled water. Hydrolysed dye preparation
The azo reactive dye chosen for the present study was Remazol Brilliant Violet 5R (Figure 1) of commercial quality (Dystar Anilinas Têxteis SA, Portugal). The dye was used without further purification. Azo reactive dyes bind to fibers by an addition mechanism under alkaline conditions and high temperature (Ganesh et al., 1994). Competing with this reaction is the hydrolysis of the dye and, as this form does not bind to fibers, it is discharged in the textile effluent. The hydrolysed dye stock solution (5 g/l) was prepared according to the dyebath conditions provided by the manufacturer. 5 g of dye were dissolved in distilled water and the pH was adjusted to 12.0±0.05 with 1 M NaOH solution. The solution was stirred for 1 hour at 80ºC. After cooling to room temperature, the solution was neutralized to pH 7.0±0.05 with 37% HCl and diluted to 1 litre with distilled water. Experimental system
The experimental system used was composed of two 1 l reactors operating in a sequencing batch mode (SBR) in a 24-hour cycle with five discrete periods: fill – 50 min, react – 21 h (mixed react – 13 h, aerated react – 8 h), settle – 1h, draw – 55 min and idle – 15 min. A feed composed of 750 mg COD/l (862 mg/l) hydrolyzed Emsize E1, 143 mg/l NH4Cl, 760 mg/l KH2PO4 and 915 mg/l Na2HPO4 was supplied to both reactors with a flow of 800 ml/day
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Figure 1 Chemical structure of Remazol Brilliant Violet 5R
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Analytical methods
Colour was measured spectrophotometrically on mixed liquor samples taken during the SBR 24-hour cycle, clarified by centrifugation. Absorbance was measured at the dye peak absorption wavelength in the visible range (λmax=560 nm) against a baseline defined by clarified samples from the dye-free reactor. Spectrophotometric analyses were performed in a Hitachi U-2000 spectrophotometer. Chemical Oxygen Demand (COD), Volatile Suspended Solids (VSS) and pH were determined according to standard procedures (American Public Health Association, 1995). Nitrate was quantified using a specific chemical analysis kit (Merck, Germany). Oxidation-Reduction Potential (ORP) was measured with an ORP electrode (platinum with silver/silver chloride as reference electrode, 3M KCl) connected to a digital pH meter. Dye degradation and metabolites formation were followed by reversed-phase HPLC (Merck-Hitachi LC-organizer, L-4250 UV-VIS detector and L-6200A intelligent pump; Hewlett Packard HP3395 integrator) using a LichroCART Purospher RP-18e column (250 mm×4 mm), (Merck, Germany) and detected at 224 nm. The eluent consisted of water and acetonitrile (gradient elution with 20–40% acetonitrile) with a flow of 1.0 ml/min.
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each. This feed was prepared in aerated tap water and the salts were analytical grade. Air was supplied by an air compressor through ceramic diffusers and additional mixing was provided by magnetic stirrers. The pumping, discharge, aeration and agitation functions were automatically controlled by a computer. Both reactors were inoculated with sludge taken from a full-scale, continuous activated sludge plant (Beirolas, Loures, Portugal) receiving mixed domestic-industrial wastewater, and acclimatized to the described feed during one month. After this acclimatization period hydrolysed dye stock solution was daily injected into one of the reactors just after the end of the fill phase, so as to produce initial concentrations of 60–100 mg dye/l.
Results and discussion SBR operation with different SRT
The SBR system was first operated with an imposed Sludge Retention Time (SRT) of 15 days (I) and then re-inoculated with fresh activated sludge for further experiments with SRT values of 10 (II) and 15 days (III).
Figure 2 Residual colour measured in the drawn effluent from the dye-fed SBR operating with SRT values of 15 days (I), 10 days (II) and 15 days (III). The reactor was re-inoculated for experiments II and III. The VSS content and initial dye concentrations in the reactor were 2.0 g VSS/l and 1.2 g VSS/l, and 90 mg/l and 60 mg/l, for SRT of 15 and 10 days, respectively
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The mixed liquor Volatile Suspended Solids (VSS) content of both reactors reached stable levels at 2.0 g VSS/l and 1.2 g VSS/l for 15 and 10 days of SRT, respectively. The temperature and pH values measured during the cycle in both SBR were always inside the 22–27ºC and 6.7–7.0 ranges, respectively. The residual colour in the drawn supernatants from the dye-fed reactor measured along 100 days of operation for each SRT experiment is represented in Figure 2. During operational period I the colour removal performance of the dye-fed SBR apparently went through an acclimatization phase during which there was an increase in the residual colour of the effluent indicating accumulation of the dye in the reactor. After a short phase of some instability, the colour removal efficiency reached a maximum, stable value around 90%, (after 40–50 days) with a fed dye concentration of 90 mg/l. After days 57 and 70 there were 4-day and 7-day periods, respectively, of imposed absence of dye in the fed substrate, after which the residual effluent colour increased only slightly before going down to lower values, again. Based on these last observations it can be assumed that the colour removal capacity of the system is not lost as a result of such short periods of lack of dye in the fed substrate. When the SRT value was lowered to 10 days (period II) colour removal levels were markedly reduced (Figures 2 and 3a). After the first week of operation the fed dye concentration was reduced from 90 to 60 mg/l so as to avoid colour accumulation in the reactor. A subsequent increase of the SRT value back to 15 days (period III) resulted in the recovery of the system’s colour and COD removal capacities and biomass concentration levels of operational period I (Figure 3). These results indicate that the adequate population for dye degradation could not be installed in the SBR when a SRT of 10 days was used. The low colour removal levels in period II could have been the results of low biomass concentration. However, some further tests were carried out with biomass grown in a parallel reactor operated in the same conditions as the dye-free, control SBR. In non-aerated decolouration runs with standardized biomass concentration, it was observed that a SRT of at least 15 days in the growth reactor was necessary to produce a consortium with high colour removal capacity (results not shown). ORP measurements–
Examples of typical ORP profiles measured during experimental periods I and II are given in Figure 4. It can be observed that in period I the dye-fed SBR experienced a reduction in the ORP values attained during the non-aerated phase, from –350 mV to lower than –400 mV, apparently as a result of the acclimatization process. This ORP change followed the colour removal improvement obtained after 40–50 days of SBR operation with this SRT
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Figure 3 Typical examples of dye (a) and COD (b) removal profiles after acclimatization for the dye-fed SBR operated at different SRT values. Cycle time started with dye injection (end of fill phase) and the aeration period was started at 13 hours of cycle time
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Figure 4 Examples of ORP profiles measured in the dye-fed and dye-free SBR reactors during operational periods I and II. The zero ORP value refers to the silver/silver chloride electrode potential
Figure 5 Examples of spectral analysis of clarified samples taken from the dye-fed SBR along day 77 of operational period III (a) and day 80 of operational period II (b). SRT values were 15 and 10 days, respectively
value (Figure 2) and agrees with the results published by Krull et al. (1998) that established a ORP value lower than –350 mV as one of the optimal conditions for azo-dye biodecolouration processes. In the dye-free, control reactor and in the dye-fed reactor operated with a SRT of 10 days (II) a reduction in the anaerobic phase ORP values was not observed (Figure 4), possibly accounting for the inefficient colour removal in the system during period II (less than 30%, Figure 3). Sample spectral analysis
To assess the possibility of colour removal resulting from adsorption of dye to the biomass, spectral analysis in the UV-visible region was performed for clarified samples taken during operational cycles. Two representative series of spectra are shown in Figure 5. For the three operational periods it was observed that the absorbance decrease was selective for the visible wavelength range, suggesting that colour removal involved structural changes in the dye molecules owing to microbial degradation and not mere physical adsorption. Also, one of the SBR cycles of period I was carried out switching the order of the aerated and the nonaerated phases and no colour removal was observed under aerobic conditions. This observation indicated that the biomass did not adsorb the dye.
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N.D. Lourenço et al. Figure 6 Colour removal performance under denitrifying conditions with an initial dye concentration of 100 mg/l. The SRT was 15 days and fed nitrate concentrations ([NO3–]o) of 45 (a) and 60 mg/l (b) were tested. The aeration started after 13 hours of cycle operation
SBR operation under denitrifying conditions
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Sodium nitrate is one of the typical salts included in dyebaths for the improvement of dye fixation to the textile fibers and the concentrations used can reach 40–100 g/l (Carliell et al., 1998). To investigate the effect of the presence of nitrate on the non-aerated biodecolouration process, additional cycles were carried out at the end of operational period I (SRT 15 days). The fed dye concentration was 100 mg/l and the feed was supplemented with nitrate concentrations of 45 and 60 mg/l. The obtained colour removal profiles and an example of nitrate removal profile are represented in Figure 6. It can be seen that colour is removed much faster when nitrate is not present in the system, which is in agreement with the results published by Carliell et al. (1998) who evaluated the interference of nitrate concentrations in the 62–620 mg/l range on azo dye biodecolouration. During the first day of operation in the presence of nitrate, differences in the colour removal profiles were not very significant, particularly for experiment (a) in Figure 6. However, on the following cycle of operation in both experiments the residual colour at the end of the anaerobic period of operation was roughly double of the values in nitrate-free runs. It can also be seen in Figure 6 (b) that nitrate was quickly removed, indicating that denitrification occurred. These results suggest that nitrate can compete with the dye for the reducing equivalents formed during the anoxic phase of the cycle and are consistent with those of Wuhrmann et al. (1980), who reported that under denitrifying conditions azo compounds were not anaerobically decoloured until all nitrate had been denitrified. This way it would not be possible to perform decolouration and denitrification in the same treatment step unless the total Hydraulic Retention Time (HRT) corresponded to the sum of the retention times required for denitrification and decolouration. It was also observed that in the dye-fed SBR operating with fed nitrate concentrations of 45 mg/l and 60 mg/l the ORP values measured during the first 2 cycle hours were close to – and –150 mV, respectively. ORP values lower than –350 mV were reached only after six hours of operation supporting the idea that nitrate is reduced preferably to the dye. The presence of nitrate did not cause significant changes in the COD removal profile.
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Analysis of metabolites
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HPLC analysis on clarified samples from the SBR mixed liquor confirmed that the azo dye was transformed and several metabolites were formed under the anaerobic conditions of the non-aerated phase. Two large peaks attributed to a benzene-based and a naphthalenebased amine (through comparison of retention times with amine standards) appeared. It was also observed that the aerobic phase introduced significant alterations in the chromatograms, mainly that the benzene-based amine was partially converted. However, the naphthalene-based amine peak remained almost unchanged throughout the aerated phase. Difficulties with the mineralization of aromatic amines under aerobic conditions after anaerobic reduction of azo dyes were also encountered by Tan et al. (1998) with the products of azo dye Mordant Orange I reduction. The suggested explanation was the absence of a suitable population of aerobic bacteria capable of metabolizing the aromatic amines. This can also be the situation in the studied system since in a 24-hour cycle only eight hours were aerobic. Conclusions
An SBR is apparently an appropriate system for azo dye biodegradation in wastewaters since it can be manipulated to provide an anaerobic phase, where azo dyes are reduced to the corresponding aromatic amines and an aerated phase where the amines can be aerobically mineralized. In the studied case colour removal of 90% was observed for a fed dye concentration of 90 mg/l and the decolouration capacity was not lost due to periods (up to at least 7 cycles) of lack of dye in the fed substrate. Based on spectral analysis of samples and on the inversion of the anaerobic-aerobic step sequence, it was concluded that colour removal was due to microbial conversion rather than adsorption. The sludge retention time (SRT) was shown to be a very important operational parameter for colour removal in the studied system. The results obtained indicated that the adequate microbial population could not be installed in the reactor with a SRT of 10 days. ORP profiles suggested that low values (under –350 mV) in the anaerobic phase are necessary for efficient colour removal to occur. The influence of the presence of nitrate in the decolouring SBR system was studied and a significant decrease in colour removal efficiency was observed after only two denitrifying cycles. These results suggest that nitrate can compete with the dye for the reducing equivalents formed during the non-aerated phase of reaction and that a combination of denitrification with colour removal in the system could only be effective if the total HRT corresponded to the sum of the retention times required for each reaction. Complete mineralization of the aromatic amines formed during the anaerobic stage was apparently not achieved in the aerated stage, indicating that further investigation should be directed to the latter. Acknowledgements
This work was financed by the European Commission Contract ENV4-CT95-0064 and FCT (Portugal, Contract PBIC/C/BIO/2027/95). The help of Dr. Westerkamp (Dystar Textilfarben, Germany) and Ir. F. van de Velde (Delft University of Technology, The Netherlands) in HPLC analyses is gratefully acknowledged. N.D. Lourenço acknowledges a PhD fellowship from FCT (Portugal). References American Public Health Association (1995), Standard Methods for the Examination of Water and Wastewater, A.D. Eaton, L.S. Clesceri and A.E. Greenberg (eds.), 19th ed., Washington D.C. Beydilli, M.I., Pavlotathis, S.G. and Tincher, W.C. (1998). Decolorization and toxicity screening of selected reactive azo dyes under methanogenic conditions. Wat. Sci. Tech., 38(4–5), 225–232.
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