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Treatment of copper wastewater by sulfate reducing bacteria in the presence of zero valent iron
International Journal of Mineral Processing 112–113 (2012) 71–76
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International Journal of Mineral Processing journal homepage: www.elsevier.com/locate/ijminpro
Treatment of copper wastewater by sulfate reducing bacteria in the presence of zero valent iron He Bai a, Yong Kang a,⁎, Hongen Quan a, Yang Han a, Ying Feng b a b
School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China School of Mechanical Engineering, Shenyang University of Chemical Engineering, Shenyang 110142, China
a r t i c l e
i n f o
Article history: Received 26 December 2011 Received in revised form 31 May 2012 Accepted 14 June 2012 Available online 21 June 2012 Keywords: Copper Sulfate reducing bacterial Zero valent iron Copper wastewater
a b s t r a c t The present study was conducted to investigate sulfate-reducing bacteria (SRB) activity and copper removal utilizing SRB enhanced by zero valent iron (Fe0) with MgSO4·7H2O and Na2SO4 as sulfate source and sodium lactate as energy resource in three distinct systems: SRB system, Fe 0 system and SRB + Fe 0 system. The SRB activity and copper removal were tested in synthetic wastewater containing 1250 mg L − 1 sulfate, 600 mg L− 1chemical oxygen demand (COD), copper (0, 5, 10, 15, 20, and 25 mg L − 1) and pH 6.0. Sulfate reductions in the SRB system and SRB + Fe 0 system were effective below 20 and 25 mg L− 1 of initial copper concentration, respectively. Copper concentrations higher than 20 mg L− 1 and 25 mg L − 1 were lethal to SRB in the SRB system and SRB + Fe0 system, respectively. The sulfate reduction rates in the SRB + Fe 0 system were increased by 559% relative to SRB system with the copper concentrations of 15 mg L − 1. The results of sulfate reduction show that the addition of Fe0 enhanced the activity of SRB. In the copper removal experiment, the overall copper removal was around 99.5% after 40 h in all three systems, whereas the SRB + Fe 0 system exhibited a better performance of copper removal than the other systems with a reaction time of 120 min. EDS (energy-dispersive spectroscopy) analysis showed that the precipitation of copper occurred in form of sulfide. The results of SRB activity and copper removal experiments demonstrated that the SRB + Fe 0 system was more capable to treat acid copper wastewater than the SRB and Fe 0 systems. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Acid mine drainage (AMD), characterized by low pH and high concentrations of heavy metals and sulfate, can severely contaminate surface and groundwater as well as soils (Akcila and Koldasb, 2006). Heavy metals such as zinc, copper, nickel and lead, are nonbiodegradable and they can accumulate in living tissues, causing various diseases and disorders; therefore, they must be removed before discharge (Ngah and Hanafiah, 2008). Several methods such as chemical precipitation, ion-exchange, reverse osmosis, electrolysis process, etc. could be applied to treat AMD for heavy metals removal. However, all of them are not good enough for AMD treatment. A major disadvantage of chemical precipitation is the need of large doses of lime and peroxide, and costly sludge disposal fees (Matlock et al., 2002). The production of concentrated brine is a significant drawback of ion-exchange as regulation of its disposal is becoming increasingly strict (Zhang et al., 2008). As for electrolysis process, it results in the high concentrations of suspended solids of the final effluent, especially composed from significant amounts of algae (Curteanu et al., 2011). The most disadvantage of reverse osmosis, besides of their high cost and increased brine volumes, is promotion of scaling with ⁎ Corresponding author. Tel./fax: + 86 22 27408813. E-mail address: [email protected] (Y. Kang). 0301-7516/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.minpro.2012.06.004
excess dosages (Malaeb and Ayoub, 2011). As a matter of fact there is an increasing interest in the biotechnological processes of AMD remediation as an alternative method for the removal of heavy metals. Biological treatment with SRB has been considered as the most promising alternative for heavy metals removal due to its low cost and high efficiency (Martins et al., 2009). The mechanisms of heavy metals removal by biological treatment include the precipitation of metal with H2S produced by SRB and biosorption onto SRB culture surface by cell wall and extracellular polysaccharides (EPS) (Pagnanelli et al., 2010). With the supplementation of organic compounds, sulfate is bioreduced to H2S under anaerobic conditions and heavy metals form stable precipitates with produced H2S. The pH of the wastewater is increased by bicarbonate during the sulfate bioreduction (Bayrakdar and Sahinkaya, 2009). In the study of Sahinkaya and Gungor, more than 99% Cu and Zn precipitation with H2S was observed in sulfatereducing up-flow reactors (UFBR) and down-flow fluidized-bed reactors (DFBR) (Sahinkaya and Gungor, 2010). The study of Teclua et al. has shown that arsenic species at concentrations of 5 mg/L or less can be removed by precipitating the metalloid out of the solution as the metal sulfide by reacting with the H2S produced by SRB (Teclua et al., 2008). The treatment of real acid mine drainage water containing sulfate and various metals was studied in an ethanol-fed sulfatereducing fluidized bed reactor, and the percent of metal removal
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was over 99.9% for Al, Co, Cu, Fe, Ni, Pb and Zn, and 94% for Mn (Sahinkaya et al., 2011). Biosorption was mainly applied to treat heavy metals wastewater with low concentration and single component. The biosorption capacity of Desulfovibrio species (a Gram-negative genus of SRB) was investigated in the removal of cadmium, zinc and copper (Pagnanelli et al., 2010; Viggi et al., 2010). Although the effectiveness of AMD bioremediation using SRB has been successfully verified during the past decade, the application of this bioremediation has been constrained due to the sensitivities of SRB to acidity and heavy metals. The selection of a remediation reagent is required to improve the living condition and enhance the activity of SRB. Fe 0 is often present as a reducing agent in the treatment of groundwater contaminated with heavy metals because of its high reduction potential, being nontoxic and inexpensive (Wilkin and McNeil, 2003). With a strong reducibility of Fe 0, water (i.e. H + in H2O) will be reduced at the iron surface, releasing hydrogen gas. The hydrogen is utilized by hydrogen-utilizing SRB as an electron donor for SO42 − reduction (Karri et al., 2005). The addition of Fe 0 has potential for sulfate reduction enhancement, pH buffering, and metals and trace elements removal. The studies of Lindsay et al. (2008) and Yao et al. (2008) demonstrated that such mixtures can support enhanced sulfate reduction. In this study, the sulfate reduction and copper removal were investigated in the SRB+ Fe0 system, the SRB system and the Fe0 system under different conditions, and the copper removal mechanisms were analyzed in different systems.
Fig. 1. The equipment of batch experiment.
HNO3 solution for 72 h to remove organic materials, rinsed with deionized water to avoid metal contamination, and dried before use. To maintain the anaerobic condition, the reactors were purged with nitrogen gas. Gas produced during the treatment process was trapped by gas collection flask.
2. Materials and methods
2.3.2. Synthetic wastewater composition The synthetic acid copper wastewater used in this study was prepared from the dissolution of copper-nitrate as heavy metal source, and hydrochloric acid and sodium hydroxide were used to adjust its pH to 6.0. Sodium lactate and sodium sulfate were added as organic carbon source and energy source respectively. Different concentrations of copper (0, 5, 10, 15, 20, and 25 mg L − 1) were contained in this synthetic wastewater.
2.1. Microorganisms
2.4. Experimental procedure
The mixed culture of SRB was initially collected from activated anaerobic sludge of a wastewater treatment plant at Hua-Bei pharmaceutical factory, Hebei, China. Sludge samples were collected from an anaerobic reaction tank and then purged with high purity nitrogen to reduce the dissolved oxygen concentration. 50 g sludge inoculum was added into 1000 mL flask with medium I (500 mg/L K2HPO4, 1000 mg/L NH4Cl, 2000 mg/L MgSO4·7H2O, 500 mg/L Na2SO4, 100 mg/L CaCl2, 1000 mg/L yeast extract, 4000 mg/L sodium lactate, 1200 mg/L ammonium ferrous sulfate), with lactate serving as the organic carbon source and sulfate as the energy source for growth. The pH of the medium was adjusted to 6 using 2 M HCl or NaOH. After purging with pure nitrogen for 5 min, the flask was sealed and incubated at 36 °C for 7 days. Then 25 mL of the mixture was inoculated into a new bottle with fresh medium I, and incubated at 36 °C for another 7 days. The enriched SRB culture was obtained by repeating the process for three times. The SRB populations were enumerated by the Most Probable Number (MPN) (Cochran, 1950). The concentration of SRB population was approximately 7.5× 106 cells mL− 1. All chemicals used in this study were of analytical grade or better and used as received.
The bioreactors were first inoculated with 10% (v/v) of the enriched SRB consortium and incubated for 7 days at batch operating conditions using medium II (500 mg L − 1 K2HPO4, 1000 mg L − 1 NH4Cl, 100 mg L − 1 CaCl2, 1250 mg L − 1 SO 2 −4). After preincubation (start of the experiment), all reactors used in batch experiments were fed with synthetic wastewater containing copper in the influent concentrations of 0, 5, 10, 15, 20, and 25 mg L − 1. The initial pH of the influent was at 6.0. The effects of copper on SRB activity and copper removal were monitored. Sulfate reduction and copper removal utilizing enhanced SRB by Fe 0 were analyzed in batch experiments in three distinct systems. The system inoculated with the enriched culture described in Section 2.1 was treated as a SRB system. The treatment system, containing the same quantity of enriched culture and 300 mg industrial grade iron powder was referred to as SRB + Fe 0 system. The system with Fe 0 but without SRB culture was treated as a Fe 0 system.
2.2. Zero valent iron The purity of iron powder was 90% containing approximately 2.5% Mn, 1.0% Si, 0.3% C, 0.1% S, 0.18% P, and 3% hydrochloric acid insoluble materials. The diameter of the particles was 255 μm. 2.3. Experimental set-up 2.3.1. Reactors The acid copper wastewater treatment was conducted in batch reactors with a working volume of 1.5 L (Fig. 1). The reactors were kept at 36 °C using an electrothermal constant magnetic stirrer and electrical control mercury thermometer. The reactor was mixed by magnetic stirrer with a speed of 400 rpm. All reactors were soaked in a 3 M
2.5. Analytical methods SO42 − was determined using a C200 Multiparameter Ion Specific Meter (Hanna Instruments, Padova, Italy). Copper was analyzed using a C200 Multiparameter Ion Specific Meter (Hanna Instruments, Padova, Italy). The detection limits for SO42 − and copper were 0.01 mg L − 1and 0.01 mg L − 1. Qualitative and quantitative analysis of precipitates was realized by EDS (energy-dispersive spectroscopy). FTIR spectra of native SRB were obtained using the Thermo Nicolet Nexus FTIR spectrophotometer (Thermo Nicolet Corporation, USA). 3. Results and discussion 3.1. SRB activity During the present work, the enriched SRB exposed to synthetic wastewater with copper was tested for activity of SRB in response
H. Bai et al. / International Journal of Mineral Processing 112–113 (2012) 71–76
to the increasing concentrations of copper. The percentage of sulfate removal was regarded as indicative of the activity of SRB. The effect of initial copper concentrations on the activity of SRB was investigated in the SRB system and the SRB + Fe 0 system under the increasing concentration of copper. The effect of initial copper concentrations on the activity of SRB was investigated in batch experiments in the presence of the increasing concentrations of copper. The percentage of sulfate removal decreases as the concentration of copper increases (Fig. 2(a)). In the absence of copper in the synthetic wastewater, the removal percentage steeply increased to 76% after 48 h, and sulfate was reduced completely at the end of the experiment (96 h). In the presence of 10 mg L − 1 of copper in the synthetic wastewater, the removal percentage of sulfate gradually increased to 8% after 96 h, 37.6% after 120 h, and 72.8% after 240 h. In case of 20 mg L − 1 of copper in the synthetic wastewater, no observable variation of sulfate concentration occurred. In this case, it was apparent that the higher the initial concentration of copper is, the stronger the inhibition to the activity of SRB. It should be noticed that the inhibited sulfate reduction (sulfate reduction b 10%) was performed at the copper concentration higher than 20 mg L − 1. The lethal copper concentration to the SRB was 20 mg L − 1 in this study, and copper tolerant SRB was able to remove copper effectively at 150 mg L − 1 copper concentration (Jalali and Balwin, 2000). Cabrera et al. (2006) reported that the SRB culture without enrichment was more sensitive to copper than that used in this study.
(a) SRB system 100
Reductuin ratio of Sulfate(%)
90 80 70 2+
-1
Cu (mg L ) 0 5 10 15 20 25
60 50 40 30 20 10 0 0
50
100
150
200
250
Time(h)
(b) SRB+Fe0 system 100
Reduction ratio of Sulfate(%)
90 80 70 2+
-1
Cu (mg L ) 0 5 10 15 20 25
60 50 40 30 20 10 0 0
20
40
60
80
100
120
140
160
180
Time (h) Fig. 2. Sulfate removal ratio at different copper concentrations in the SRB system and SRB + Fe0 system. (a) SRB system. (b) SRB + Fe0 system.
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Bacterial sulfate reduction percentage with increasing initial concentration of copper in the SRB + Fe 0 system is shown in Fig. 2(b). In the absence of copper in the synthetic wastewater, the percentage of sulfate reduction was to 84.9% after 48 h, and the sulfate was completely reduced within 96 h. The percentage of sulfate reduction was 4.76% after 48 h when initial copper concentration was 10 mg L− 1, while the sulfate was entirely reduced at the end of the experiment (120 h). At the highest copper concentration (25 mg L− 1), the percentage of sulfate reduction increased gradually to about 13.39% after 96 h, then reached 55.91% after 120 h. The removal percentage of sulfate increased to 83.46% around 168 h which was much higher compared with 0.8% in the SRB system. It is reasonable to conclude that the lethal concentration of copper to SRB in SRB + Fe 0 system was higher than 25 mg L − 1, much higher than that in the SRB system. The lag times of the SRB culture during the 240 h experiments with different initial copper concentrations are shown in Table 1. The lag time increased as initial concentrations of copper shifted from 0 to 25 mg L − 1, indicating that the growth of SRB culture was inhibited strongly with the increase of initial copper concentrations. The lag times in SRB system were longer than those in SRB + Fe0 system. For example, the lag time in the SRB system and SRB+ Fe 0 system was 110–120 and 55–60 h at 15 mg L − 1 of copper, respectively. It is noticed that the lag time was more than 240 h in the SRB system at 20 mg L − 1 of copper, which was consistent with the results of removal ratio of sulfate at this copper concentration. The percentage of sulfate reduction and sulfate reduction rate was found to be correlated with copper concentration. The percentage of sulfate reduction was correlated to the copper concentration as shown in Fig. 3(a). It shows the profiles of sulfate reduction ratio for samples containing various initial copper concentrations, revealing that the percentage of sulfate reduction in the SRB + Fe 0 system was higher than that in the SRB system. For example, the percentage of sulfate reduction in SRB system was 2.4% relative to 88.8% in the SRB + Fe 0 system, when the copper concentration was 20 mg L − 1. Comparing the percentage of sulfate reduction between the SRB system and the SRB + Fe 0 system, it is apparent that the SRB + Fe 0 system performed elevated capability of sulfate reduction. The rate of sulfate reduction versus copper concentration was plotted in Fig. 3(b). Compared to the blank treatment in the SRB system, the reduction rates were decreased gradually by 78.9%, 98.4%, and 100% while copper concentrations were 10, 20, 25 mg L − 1, indicating that the sulfate reduction rate was constrained intensively by initial copper concentration. In fact, the average sulfate reduction rates were 40.53 L − 1 h − 1 and 20.1 mg L − 1 h − 1 in the SRB system and SRB + Fe 0 system, respectively. SRB is sensitive to heavy metals which result in the denaturation and inactivation of enzymes and the disruption of cell organelles and membrane integrity (Alexandrinoa et al., 2011). The inhibition of inner enzyme synthesis leads to a limited sulfate reduction ratio at high initial copper concentration in the SRB system, which resulted in the decrease of sulfate reduction and SRB activity. Fe 0 was a very important trace element for bacterial metabolism, since it was integrated into iron–sulfur clusters that serve to bind substrates into enzyme active sites. In SRB + Fe 0 system, the SRB culture exhibited a high rate of sulfate reduction in the presence of low concentration of Fe 0. The sulfate reduction rate in the SRB + Fe 0 system was
Table 1 Lag times of sulfate reducing at different copper concentrations in the SRB system and SRB + Fe0 system. Cu2+ (mg L− 1)
SRB (h)
SRB + Fe0 (h)
Cu2+ (mg L− 1)
SRB (h)
SRB + Fe0 (h)
0 5 10
30–35 50–55 70–75
30–35 40–45 40–45
15 20 25
110–120 >240 >240
55–60 75–80 85–90
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(a) Results of sulfate reduction ratio at different copper concentrations in SRB system and SRB+Fe0 system
(a) Change of Copperwith time at different copper concentrations in SRB system and SRB+Fe0 system
0
SRB+Fe system SRB system
25
Cu
2+
-1
(mg L ) 5
80
10
20
Cu2+ (mg L-1)
Sulfate reduction ratio (%)
100
60
40
15 20
15
25 10
20 5 0 0
5
10
15
20
25 0
Cu2+ (mg L-1)
0
40
copper concentrations in SRB system and SRB+Fe0 system 0
SRB+Fe system SRB system
120
160
200
(b) Compare of copper removal ratio at copper concentration of 25 mgL-1in SRB system, Fe0 system,and SRB+Fe0 system 100 90
80
Removal ratio of cooper (%)
Sufate reduction rate (mg L-1 h-1)
100
80
Time (hour)
(b) Relation of sulfate reducing rates with different
60
40
20
0 0
5
10
15
20
25
Cu2+ (mg L-1)
80 70 60 50 40
0
30
Fe SRB
20
SRB+Fe
0
10 0
Fig. 3. Effect of copper on the sulfate removal ratios and rates of sulfate reduction with different copper concentrations in the SRB system and SRB + Fe0 system. (a) Results of sulfate reduction ratio at different copper concentrations in the SRB system and SRB + Fe0 system. (b) Relation of sulfate reducing rates with different copper concentrations in the SRB system and SRB + Fe0 system.
increased by 559% relative to the SRB system with the copper concentration of 15 mg L − 1. It is confirmed that the supplementation of Fe 0 in the SRB culture accelerates the removal of sulfate effectively.
0
30
60
90
120
Time (minute) Fig. 4. Removal of copper with time in the SRB system, Fe0 system, and SRB + Fe0 system. (a) Change of copper with time at different copper concentrations in the SRB system and SRB + Fe0 system. (b) Comparison of copper removal ratio at copper concentration of 25 mg L− 1 in the SRB system, Fe0 system, and SRB + Fe0 system.
3.2. Copper removal
However, at the end of the experiment (120 min), the final removal ratios in the SRB system and SRB + Fe 0 system were similar; much higher than that in Fe 0 system.
3.2.1. Removal of copper In order to investigate the removal mechanism of copper, Fe 0 mentioned in Section 2.2 was added into the treatment reactor without SRB culture, which was regarded as Fe 0 system. As mentioned, during the biodegrading procedure, the sulfate and lactate were contained in the synthetic wastewater at pH 6.0 and at 36 °C. No apparent differences in the copper removal were observed in all systems (Fig. 4(a)), indicating that copper removal ratio below 0.1 mg L − 1 was achieved after 48 h at all copper concentrations in all systems. The capability and mechanism of copper removal were analyzed within 120 min at copper concentration of 25 mg L − 1. Fig. 4(b) shows that copper was removed faster in the SRB + Fe 0 system than in the SRB system and Fe 0 system. With an initial copper concentration of 25 mg L − 1 in the SRB + Fe 0 system, copper removal ratio increased from 54% to 96% after 45 min, whereas in the SRB system and Fe 0 system, copper removal ratio were only 83% and 44%.
3.2.2. Chemical composition of the precipitate In the present study, the mechanism for copper removal was apparently precipitation with sulfide. The chemical composition of the precipitate from the SRB + Fe 0 system was qualitatively analyzed with EDS at the end of the experiments. The chemical elements content of the precipitate was confirmed to be copper sulfide by the EDS spectrum shown in Fig. 5. It was revealed in Table 2 that the chemical composition of the precipitates contained large amounts of sulfur (59.81%), oxygen (28.24%), and a lesser amount of copper (5.86%), Fe, Ca, and P. In this case, the amount of sulfur was abundant to copper resulting in the complete removal of copper by means of apparent precipitation with sulfide. According to the result of XRD (data was not shown), the precipitation contains large amounts of Cu7S4, a little amount of Cu 0, CuFeS2, and Cu9Fe9S16, and insignificant amounts of oxide phases.
H. Bai et al. / International Journal of Mineral Processing 112–113 (2012) 71–76
75
621.63 534.18 1234.30 1081.84
1536.76
1453.62 1400.73
2960.27 2926.64 2856.28
80
1651.11
3294.04
Transmittance (%)
2360.53
100
Fig. 5. EDS analysis of metal precipitation sample.
In addition to precipitation with sulfide, copper might be removed through sorption on the biomass (Jalali and Balwin, 2000). In fact, SRB cells played an additional, important role in the facilitation of metal precipitation. 3.2.3. Removal mechanism The mechanisms of copper removal in the study including the effect of SRB culture and Fe 0 are rather complex. These mechanisms are integral components of biosorption, sulfide precipitation, and reductive precipitation (Gadd, 2000). The sulfate reduction ratios in the SRB system and SRB + Fe 0 system maintained at high level at low initial copper concentration indicate an efficient capability of sulfate reduction. Almost the entire copper was removed by means of precipitation with abundant sulfide produced by sulfate reduction, which was confirmed by the results of EDS analysis of the precipitate samples. It was concluded that sulfide precipitation contributes to copper removal at low initial copper concentration in the SRB system and SRB + Fe 0 system. However, the removal ratio of copper was still high with the sulfate reduction inhibited partly or completely at the high initial copper concentration in the SRB system. Copper removal to more than 95% was achieved after 216 h in the presence of 4% sulfate reduction at 25 mg L − 1 initial concentration of copper. The sulfide produced from sulfate reduction by SRB was insufficient to precipitate with copper at the high initial copper concentration. In the SRB system, it was found that the bacteria directly bind the metals in their cell walls and extracellular polymeric substances (Wang and Chen, 2006). Biosorption is either metabolism independent such as physical or chemical sorption onto the cell wall, or metabolism related such as transport and extracellular precipitation by metabolites (Gadd, 2000). Considering a high metal-reducing capability of SRB, it is suggested that SRB has a significant capacity of metal biosorption. To confirm the type of functional groups of SRB, FTIR spectra of native SRB were obtained by using a FTIR spectrophotometer. As shown in Fig. 6, The FTIR spectra of biomass indicate the presence of amino, carboxylic, hydroxyl and carbonyl groups. Display of strong broad \OH stretch carboxylic bands in the region 3294 cm − 1 and carboxylic/ phenolic stretching bands in the region of 2926 cm− 1 was observed. The peaks in the region 1651 cm− 1 were attributed to C_N, C_C and C_O stretch whereas the peaks appearing in the region 1536 cm− 1 might represent quinine O\H bond. Now the peaks appearing in the region 1356, 1081 and 1453 cm− 1 represent N\H bending, \CH3 Table 2 Element contents of sediment by EDS analysis of metal precipitation sample. Element
Composition (%)
Element
Composition (%)
O P S
28.24 1.06 59.81
Ca Fe Cu
0.78 4.25 5.86
60 4000
3500
3000
2500
2000
1500
1000
500
Wavenumber (cm-1) Fig. 6. FTIR spectra of function groups in the surface of SRB culture.
wagging, and the bending of \CH3, respectively. The biomass showed bands around 1400 and 1234 cm− 1 representing the ionized carboxylate group (COO\). The effect of Fe 0 on copper removal was studied in Fe 0 system in the absence of SRB. The copper removal was 62.2% after 120 min and reached 95% at the end of the experiment (160 h). Because standard potential of copper was more positive than iron, the reductive precipitation was predominant removal mechanism in the Fe 0 system (Li and Zhang, 2007). When a metal is converted to a lower redox state, its mobility and toxicity can be reduced. It is proposed that, for wastewater with high copper concentration, Fe 0 is an efficient reluctant for reduction of cooper. At low initial concentration of copper, the sulfate reduction ratio and copper removal were similar in the SRB system and SRB + Fe 0 system, indicating that the sulfide precipitation contributes to the copper removal. With the increasing copper concentration, sulfate reduction ratio in the SRB + Fe 0 system performed a higher level than that in the SRB system for enhanced activity of SRB by Fe 0. When the initial concentration of copper was 25 mg L − 1, the sulfate reduction ratio was still 83.46% after 160 h in the SRB + Fe 0 system. Considering the results of sulfate reduction and copper removal, it is assumed that reductive precipitation and biosorption play important roles in copper removal since the activity of SRB was constrained when copper concentration was above 25 mg L − 1. In this case, the cooperation of sulfate precipitation, biosorption and reductive precipitation occurred to remove copper in the SRB + Fe 0 system. For all copper concentrations, SRB augmented by Fe 0 was capable of removing copper completely in the SRB + Fe 0 system with sulfide precipitation and biosorption and reductive precipitation. It was demonstrated that performance of the SRB + Fe 0 system was significantly enhanced compared with the SRB and Fe 0 systems. 4. Conclusions The activity of SRB enhanced by Fe 0 played an important role in copper removal in the presence of high concentrations of copper (20 mg L − 1 and 25 mg L − 1). The 84.36% of sulfate reduction ratio in SRB + Fe 0 system was much higher than the 5.6% in the SRB system after 216 h at 25 mg L − 1of copper. The sulfate reduction rate at 82 mg L − 1 h − 1 was achieved in the presence of 25 mg L − 1 of copper in SRB + Fe 0 system, whereas only 5.6 mg L − 1 h − 1 of sulfate reduction rate was observed in the SRB system. Similar capability of copper removal was observed in all systems, and the overall copper removal was around 99.5% after 40 h in all three systems, whereas the SRB + Fe 0 system exhibited a better performance of copper removal than the other systems within 120 min. In the copper removal experiment, EDS analysis indicated the
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formation of a dominating crystalline structure that was mainly composed of Cu7S4. Sulfide precipitation and biosorption contribute to the copper removal in the SRB system. In contrast, reductive precipitation played an important role in the removal of copper in the Fe 0 system. The removal of copper was most effective in the SRB + Fe 0 system due to the cooperation of sulfide precipitation, biosorption and reductive precipitation. Performance of the SRB + Fe 0 system was significantly enhanced compared with the SRB and Fe 0 systems. Acknowledgments The authors are grateful for the financial support of National Nature Science Foundation of China (Project Nos. 21077075 and 20106014). References Akcila, A., Koldasb, S., 2006. Acid Mine Drainage (AMD): causes, treatment and case studies. J. Clean. Prod. 14, 1139–1145. Bayrakdar, A., Sahinkaya, E., 2009. Performance of sulfidogenic anaerobic baffled reactor (ABR) treating acidic and zinc-containing wastewater. Bioresour. Technol. 100, 4354–4360. Viggi, C., Pagnanelli, F., Cibati, A., Uccelletti, D., Palleschi, C., Toro, L., 2010. Biotreatment and bioassessment of heavy metal removal by sulphate reducing bacteria in fixed bed reactors. Water Res. 44, 151–158. Teclua, D., Tivchev, G., Laing, M., Wallis, M., 2008. Bioremoval of arsenic species from contaminated waters by sulphate reducing bacteria. Water Res. 42, 4885–4893. Sahinkaya, E., Gunes, F.M., Ucar, D., Kaksonen, A.H., 2011. Sulfidogenic fluidized bed treatment of real acid mine drainage water. Bioresour. Technol. 102, 683–689. Sahinkaya, E., Gungor, M., 2010. Comparison of sulfidogenic up-flow and down-flow fluidized-bed reactors for the biotreatment of acidic metal-containing wastewater. Bioresour. Technol. 101, 9508–9514. Pagnanelli, F., Viggi, C.C., Toro, L., 2010. Isolation and quantification of cadmium removal mechanisms in batch reactors inoculated by sulphate reducing bacteria: biosorption versus bioprecipitation. Bioresour. Technol. 101, 2981–2987. Cabrera, G., Pérez, R., Gómez, J.M., Ábalos, A., Cantero, D., 2006. Toxic effects of dissolved heavy metals on Desulfovibrio vulgaris and Desulfovibrio sp. strains. J. Hazard. Mater. 135, 40–46. Gadd, G.M., 2000. Bioremedial potential of microbial mechanisms of metal mobilization and immobilization. Curr. Opin. Biotechnol. 11, 271–279.
Wang, J., Chen, C., 2006. Biosorption of heavy metals by Saccharomyces cerevisiae: a review. Biotechnol. Adv. 24, 427–451. Jalali, K., Balwin, S.A., 2000. The role of sulfate-reducing bacteria in copper removal from aqueous sulfate solutions. Water Res. 34, 797–806. Malaeb, L., Ayoub, G.M., 2011. Reverse osmosis technology for water treatment: state of the art review. Desalination 267, 1–8. Zhang, L.H., De Schryver, P., De Gussemea, B., De Muynck, W., Boon, N., Verstraete, W., 2008. Chemical and biological technologies for hydrogen sulfide emission control in sewer systems: a review. Water Res. 42, 1–12. Martins, M., Faleiro, M.L., Barros, R.J., Raquel Veríssimo, A., Barreiros, M.A., Clara Costa, M., 2009. Characterization and activity studies of highly heavy metal resistant sulphate-reducing bacteria to be used in acid mine drainage decontamination. J. Hazard. Mater. 166, 706–713. Matlock, M., Howerton, B.S., Atwood, D.A., 2002. Chemical precipitation of heavy metals from acid mine drainage. Water Res. 36, 4757–4764. Alexandrinoa, M., Macíasb, F., Costaa, R., Gomesc, N.C.M., Canárioa, A.V.M., Costa, M.C., 2011. A bacterial consortium isolated from an Icelandic fumarole displays exceptionally high levels of sulfate reduction and metals resistance. J. Hazard. Mater. 137, 162–370. Lindsay, M.B.J., Ptacek, C.J., Blowes, D.W., Douglas Gould, W., 2008. Zero-valent iron and organic carbon mixtures for remediation of acid mine drainage: batch experiments. Appl. Geochem. 23, 2214–2225. Wilkin, R.T., McNeil, M.S., 2003. Laboratory evaluation of zero-valent iron to treat water impacted by acid mine drainage. Chemosphere 53, 715–725. Curteanu, S., Piuleac, C.G., Godini, K., Azaryan, G., 2011. Modeling of electrolysis process in wastewater treatment using different types of neural networks. Chem. Eng. J. 172, 267–276. Karri, S., Alvarez, R.S., Field, J.A., 2005. Zero-valent iron as an electron donor for methanogenesis and sulfate reduction in anaerobic sludge. Bioresour. Technol. 92, 810–819. Cochran, W.G., 1950. Estimation of bacterial densities by means of the most probable number. Biometrics 6, 105–116. Ngah, W.S.W., Hanafiah, M.A.K.M., 2008. Removal of heavy metal ions from wastewater by chemically modified plant wastes as adsorbents: a review. Bioresour. Technol. 99, 3935–3948. Li, X., Zhang, W., 2007. Sequestration of metal cations with zerovalent iron nanoparticles — a study with high resolution X-ray photoelectron spectroscopy (HR-XPS). J. Phys. Chem. C 111, 6939–6946. Yao, X., Kang, Y., Lee, D., Feng, Y., 2008. Bioaugmented sulfate reduction using enriched anaerobic microflora in the presence of zero valent iron. Chemosphere 7, 1436–1441.
Contents lists available at SciVerse ScienceDirect
International Journal of Mineral Processing journal homepage: www.elsevier.com/locate/ijminpro
Treatment of copper wastewater by sulfate reducing bacteria in the presence of zero valent iron He Bai a, Yong Kang a,⁎, Hongen Quan a, Yang Han a, Ying Feng b a b
School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China School of Mechanical Engineering, Shenyang University of Chemical Engineering, Shenyang 110142, China
a r t i c l e
i n f o
Article history: Received 26 December 2011 Received in revised form 31 May 2012 Accepted 14 June 2012 Available online 21 June 2012 Keywords: Copper Sulfate reducing bacterial Zero valent iron Copper wastewater
a b s t r a c t The present study was conducted to investigate sulfate-reducing bacteria (SRB) activity and copper removal utilizing SRB enhanced by zero valent iron (Fe0) with MgSO4·7H2O and Na2SO4 as sulfate source and sodium lactate as energy resource in three distinct systems: SRB system, Fe 0 system and SRB + Fe 0 system. The SRB activity and copper removal were tested in synthetic wastewater containing 1250 mg L − 1 sulfate, 600 mg L− 1chemical oxygen demand (COD), copper (0, 5, 10, 15, 20, and 25 mg L − 1) and pH 6.0. Sulfate reductions in the SRB system and SRB + Fe 0 system were effective below 20 and 25 mg L− 1 of initial copper concentration, respectively. Copper concentrations higher than 20 mg L− 1 and 25 mg L − 1 were lethal to SRB in the SRB system and SRB + Fe0 system, respectively. The sulfate reduction rates in the SRB + Fe 0 system were increased by 559% relative to SRB system with the copper concentrations of 15 mg L − 1. The results of sulfate reduction show that the addition of Fe0 enhanced the activity of SRB. In the copper removal experiment, the overall copper removal was around 99.5% after 40 h in all three systems, whereas the SRB + Fe 0 system exhibited a better performance of copper removal than the other systems with a reaction time of 120 min. EDS (energy-dispersive spectroscopy) analysis showed that the precipitation of copper occurred in form of sulfide. The results of SRB activity and copper removal experiments demonstrated that the SRB + Fe 0 system was more capable to treat acid copper wastewater than the SRB and Fe 0 systems. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Acid mine drainage (AMD), characterized by low pH and high concentrations of heavy metals and sulfate, can severely contaminate surface and groundwater as well as soils (Akcila and Koldasb, 2006). Heavy metals such as zinc, copper, nickel and lead, are nonbiodegradable and they can accumulate in living tissues, causing various diseases and disorders; therefore, they must be removed before discharge (Ngah and Hanafiah, 2008). Several methods such as chemical precipitation, ion-exchange, reverse osmosis, electrolysis process, etc. could be applied to treat AMD for heavy metals removal. However, all of them are not good enough for AMD treatment. A major disadvantage of chemical precipitation is the need of large doses of lime and peroxide, and costly sludge disposal fees (Matlock et al., 2002). The production of concentrated brine is a significant drawback of ion-exchange as regulation of its disposal is becoming increasingly strict (Zhang et al., 2008). As for electrolysis process, it results in the high concentrations of suspended solids of the final effluent, especially composed from significant amounts of algae (Curteanu et al., 2011). The most disadvantage of reverse osmosis, besides of their high cost and increased brine volumes, is promotion of scaling with ⁎ Corresponding author. Tel./fax: + 86 22 27408813. E-mail address: [email protected] (Y. Kang). 0301-7516/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.minpro.2012.06.004
excess dosages (Malaeb and Ayoub, 2011). As a matter of fact there is an increasing interest in the biotechnological processes of AMD remediation as an alternative method for the removal of heavy metals. Biological treatment with SRB has been considered as the most promising alternative for heavy metals removal due to its low cost and high efficiency (Martins et al., 2009). The mechanisms of heavy metals removal by biological treatment include the precipitation of metal with H2S produced by SRB and biosorption onto SRB culture surface by cell wall and extracellular polysaccharides (EPS) (Pagnanelli et al., 2010). With the supplementation of organic compounds, sulfate is bioreduced to H2S under anaerobic conditions and heavy metals form stable precipitates with produced H2S. The pH of the wastewater is increased by bicarbonate during the sulfate bioreduction (Bayrakdar and Sahinkaya, 2009). In the study of Sahinkaya and Gungor, more than 99% Cu and Zn precipitation with H2S was observed in sulfatereducing up-flow reactors (UFBR) and down-flow fluidized-bed reactors (DFBR) (Sahinkaya and Gungor, 2010). The study of Teclua et al. has shown that arsenic species at concentrations of 5 mg/L or less can be removed by precipitating the metalloid out of the solution as the metal sulfide by reacting with the H2S produced by SRB (Teclua et al., 2008). The treatment of real acid mine drainage water containing sulfate and various metals was studied in an ethanol-fed sulfatereducing fluidized bed reactor, and the percent of metal removal
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was over 99.9% for Al, Co, Cu, Fe, Ni, Pb and Zn, and 94% for Mn (Sahinkaya et al., 2011). Biosorption was mainly applied to treat heavy metals wastewater with low concentration and single component. The biosorption capacity of Desulfovibrio species (a Gram-negative genus of SRB) was investigated in the removal of cadmium, zinc and copper (Pagnanelli et al., 2010; Viggi et al., 2010). Although the effectiveness of AMD bioremediation using SRB has been successfully verified during the past decade, the application of this bioremediation has been constrained due to the sensitivities of SRB to acidity and heavy metals. The selection of a remediation reagent is required to improve the living condition and enhance the activity of SRB. Fe 0 is often present as a reducing agent in the treatment of groundwater contaminated with heavy metals because of its high reduction potential, being nontoxic and inexpensive (Wilkin and McNeil, 2003). With a strong reducibility of Fe 0, water (i.e. H + in H2O) will be reduced at the iron surface, releasing hydrogen gas. The hydrogen is utilized by hydrogen-utilizing SRB as an electron donor for SO42 − reduction (Karri et al., 2005). The addition of Fe 0 has potential for sulfate reduction enhancement, pH buffering, and metals and trace elements removal. The studies of Lindsay et al. (2008) and Yao et al. (2008) demonstrated that such mixtures can support enhanced sulfate reduction. In this study, the sulfate reduction and copper removal were investigated in the SRB+ Fe0 system, the SRB system and the Fe0 system under different conditions, and the copper removal mechanisms were analyzed in different systems.
Fig. 1. The equipment of batch experiment.
HNO3 solution for 72 h to remove organic materials, rinsed with deionized water to avoid metal contamination, and dried before use. To maintain the anaerobic condition, the reactors were purged with nitrogen gas. Gas produced during the treatment process was trapped by gas collection flask.
2. Materials and methods
2.3.2. Synthetic wastewater composition The synthetic acid copper wastewater used in this study was prepared from the dissolution of copper-nitrate as heavy metal source, and hydrochloric acid and sodium hydroxide were used to adjust its pH to 6.0. Sodium lactate and sodium sulfate were added as organic carbon source and energy source respectively. Different concentrations of copper (0, 5, 10, 15, 20, and 25 mg L − 1) were contained in this synthetic wastewater.
2.1. Microorganisms
2.4. Experimental procedure
The mixed culture of SRB was initially collected from activated anaerobic sludge of a wastewater treatment plant at Hua-Bei pharmaceutical factory, Hebei, China. Sludge samples were collected from an anaerobic reaction tank and then purged with high purity nitrogen to reduce the dissolved oxygen concentration. 50 g sludge inoculum was added into 1000 mL flask with medium I (500 mg/L K2HPO4, 1000 mg/L NH4Cl, 2000 mg/L MgSO4·7H2O, 500 mg/L Na2SO4, 100 mg/L CaCl2, 1000 mg/L yeast extract, 4000 mg/L sodium lactate, 1200 mg/L ammonium ferrous sulfate), with lactate serving as the organic carbon source and sulfate as the energy source for growth. The pH of the medium was adjusted to 6 using 2 M HCl or NaOH. After purging with pure nitrogen for 5 min, the flask was sealed and incubated at 36 °C for 7 days. Then 25 mL of the mixture was inoculated into a new bottle with fresh medium I, and incubated at 36 °C for another 7 days. The enriched SRB culture was obtained by repeating the process for three times. The SRB populations were enumerated by the Most Probable Number (MPN) (Cochran, 1950). The concentration of SRB population was approximately 7.5× 106 cells mL− 1. All chemicals used in this study were of analytical grade or better and used as received.
The bioreactors were first inoculated with 10% (v/v) of the enriched SRB consortium and incubated for 7 days at batch operating conditions using medium II (500 mg L − 1 K2HPO4, 1000 mg L − 1 NH4Cl, 100 mg L − 1 CaCl2, 1250 mg L − 1 SO 2 −4). After preincubation (start of the experiment), all reactors used in batch experiments were fed with synthetic wastewater containing copper in the influent concentrations of 0, 5, 10, 15, 20, and 25 mg L − 1. The initial pH of the influent was at 6.0. The effects of copper on SRB activity and copper removal were monitored. Sulfate reduction and copper removal utilizing enhanced SRB by Fe 0 were analyzed in batch experiments in three distinct systems. The system inoculated with the enriched culture described in Section 2.1 was treated as a SRB system. The treatment system, containing the same quantity of enriched culture and 300 mg industrial grade iron powder was referred to as SRB + Fe 0 system. The system with Fe 0 but without SRB culture was treated as a Fe 0 system.
2.2. Zero valent iron The purity of iron powder was 90% containing approximately 2.5% Mn, 1.0% Si, 0.3% C, 0.1% S, 0.18% P, and 3% hydrochloric acid insoluble materials. The diameter of the particles was 255 μm. 2.3. Experimental set-up 2.3.1. Reactors The acid copper wastewater treatment was conducted in batch reactors with a working volume of 1.5 L (Fig. 1). The reactors were kept at 36 °C using an electrothermal constant magnetic stirrer and electrical control mercury thermometer. The reactor was mixed by magnetic stirrer with a speed of 400 rpm. All reactors were soaked in a 3 M
2.5. Analytical methods SO42 − was determined using a C200 Multiparameter Ion Specific Meter (Hanna Instruments, Padova, Italy). Copper was analyzed using a C200 Multiparameter Ion Specific Meter (Hanna Instruments, Padova, Italy). The detection limits for SO42 − and copper were 0.01 mg L − 1and 0.01 mg L − 1. Qualitative and quantitative analysis of precipitates was realized by EDS (energy-dispersive spectroscopy). FTIR spectra of native SRB were obtained using the Thermo Nicolet Nexus FTIR spectrophotometer (Thermo Nicolet Corporation, USA). 3. Results and discussion 3.1. SRB activity During the present work, the enriched SRB exposed to synthetic wastewater with copper was tested for activity of SRB in response
H. Bai et al. / International Journal of Mineral Processing 112–113 (2012) 71–76
to the increasing concentrations of copper. The percentage of sulfate removal was regarded as indicative of the activity of SRB. The effect of initial copper concentrations on the activity of SRB was investigated in the SRB system and the SRB + Fe 0 system under the increasing concentration of copper. The effect of initial copper concentrations on the activity of SRB was investigated in batch experiments in the presence of the increasing concentrations of copper. The percentage of sulfate removal decreases as the concentration of copper increases (Fig. 2(a)). In the absence of copper in the synthetic wastewater, the removal percentage steeply increased to 76% after 48 h, and sulfate was reduced completely at the end of the experiment (96 h). In the presence of 10 mg L − 1 of copper in the synthetic wastewater, the removal percentage of sulfate gradually increased to 8% after 96 h, 37.6% after 120 h, and 72.8% after 240 h. In case of 20 mg L − 1 of copper in the synthetic wastewater, no observable variation of sulfate concentration occurred. In this case, it was apparent that the higher the initial concentration of copper is, the stronger the inhibition to the activity of SRB. It should be noticed that the inhibited sulfate reduction (sulfate reduction b 10%) was performed at the copper concentration higher than 20 mg L − 1. The lethal copper concentration to the SRB was 20 mg L − 1 in this study, and copper tolerant SRB was able to remove copper effectively at 150 mg L − 1 copper concentration (Jalali and Balwin, 2000). Cabrera et al. (2006) reported that the SRB culture without enrichment was more sensitive to copper than that used in this study.
(a) SRB system 100
Reductuin ratio of Sulfate(%)
90 80 70 2+
-1
Cu (mg L ) 0 5 10 15 20 25
60 50 40 30 20 10 0 0
50
100
150
200
250
Time(h)
(b) SRB+Fe0 system 100
Reduction ratio of Sulfate(%)
90 80 70 2+
-1
Cu (mg L ) 0 5 10 15 20 25
60 50 40 30 20 10 0 0
20
40
60
80
100
120
140
160
180
Time (h) Fig. 2. Sulfate removal ratio at different copper concentrations in the SRB system and SRB + Fe0 system. (a) SRB system. (b) SRB + Fe0 system.
73
Bacterial sulfate reduction percentage with increasing initial concentration of copper in the SRB + Fe 0 system is shown in Fig. 2(b). In the absence of copper in the synthetic wastewater, the percentage of sulfate reduction was to 84.9% after 48 h, and the sulfate was completely reduced within 96 h. The percentage of sulfate reduction was 4.76% after 48 h when initial copper concentration was 10 mg L− 1, while the sulfate was entirely reduced at the end of the experiment (120 h). At the highest copper concentration (25 mg L− 1), the percentage of sulfate reduction increased gradually to about 13.39% after 96 h, then reached 55.91% after 120 h. The removal percentage of sulfate increased to 83.46% around 168 h which was much higher compared with 0.8% in the SRB system. It is reasonable to conclude that the lethal concentration of copper to SRB in SRB + Fe 0 system was higher than 25 mg L − 1, much higher than that in the SRB system. The lag times of the SRB culture during the 240 h experiments with different initial copper concentrations are shown in Table 1. The lag time increased as initial concentrations of copper shifted from 0 to 25 mg L − 1, indicating that the growth of SRB culture was inhibited strongly with the increase of initial copper concentrations. The lag times in SRB system were longer than those in SRB + Fe0 system. For example, the lag time in the SRB system and SRB+ Fe 0 system was 110–120 and 55–60 h at 15 mg L − 1 of copper, respectively. It is noticed that the lag time was more than 240 h in the SRB system at 20 mg L − 1 of copper, which was consistent with the results of removal ratio of sulfate at this copper concentration. The percentage of sulfate reduction and sulfate reduction rate was found to be correlated with copper concentration. The percentage of sulfate reduction was correlated to the copper concentration as shown in Fig. 3(a). It shows the profiles of sulfate reduction ratio for samples containing various initial copper concentrations, revealing that the percentage of sulfate reduction in the SRB + Fe 0 system was higher than that in the SRB system. For example, the percentage of sulfate reduction in SRB system was 2.4% relative to 88.8% in the SRB + Fe 0 system, when the copper concentration was 20 mg L − 1. Comparing the percentage of sulfate reduction between the SRB system and the SRB + Fe 0 system, it is apparent that the SRB + Fe 0 system performed elevated capability of sulfate reduction. The rate of sulfate reduction versus copper concentration was plotted in Fig. 3(b). Compared to the blank treatment in the SRB system, the reduction rates were decreased gradually by 78.9%, 98.4%, and 100% while copper concentrations were 10, 20, 25 mg L − 1, indicating that the sulfate reduction rate was constrained intensively by initial copper concentration. In fact, the average sulfate reduction rates were 40.53 L − 1 h − 1 and 20.1 mg L − 1 h − 1 in the SRB system and SRB + Fe 0 system, respectively. SRB is sensitive to heavy metals which result in the denaturation and inactivation of enzymes and the disruption of cell organelles and membrane integrity (Alexandrinoa et al., 2011). The inhibition of inner enzyme synthesis leads to a limited sulfate reduction ratio at high initial copper concentration in the SRB system, which resulted in the decrease of sulfate reduction and SRB activity. Fe 0 was a very important trace element for bacterial metabolism, since it was integrated into iron–sulfur clusters that serve to bind substrates into enzyme active sites. In SRB + Fe 0 system, the SRB culture exhibited a high rate of sulfate reduction in the presence of low concentration of Fe 0. The sulfate reduction rate in the SRB + Fe 0 system was
Table 1 Lag times of sulfate reducing at different copper concentrations in the SRB system and SRB + Fe0 system. Cu2+ (mg L− 1)
SRB (h)
SRB + Fe0 (h)
Cu2+ (mg L− 1)
SRB (h)
SRB + Fe0 (h)
0 5 10
30–35 50–55 70–75
30–35 40–45 40–45
15 20 25
110–120 >240 >240
55–60 75–80 85–90
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(a) Results of sulfate reduction ratio at different copper concentrations in SRB system and SRB+Fe0 system
(a) Change of Copperwith time at different copper concentrations in SRB system and SRB+Fe0 system
0
SRB+Fe system SRB system
25
Cu
2+
-1
(mg L ) 5
80
10
20
Cu2+ (mg L-1)
Sulfate reduction ratio (%)
100
60
40
15 20
15
25 10
20 5 0 0
5
10
15
20
25 0
Cu2+ (mg L-1)
0
40
copper concentrations in SRB system and SRB+Fe0 system 0
SRB+Fe system SRB system
120
160
200
(b) Compare of copper removal ratio at copper concentration of 25 mgL-1in SRB system, Fe0 system,and SRB+Fe0 system 100 90
80
Removal ratio of cooper (%)
Sufate reduction rate (mg L-1 h-1)
100
80
Time (hour)
(b) Relation of sulfate reducing rates with different
60
40
20
0 0
5
10
15
20
25
Cu2+ (mg L-1)
80 70 60 50 40
0
30
Fe SRB
20
SRB+Fe
0
10 0
Fig. 3. Effect of copper on the sulfate removal ratios and rates of sulfate reduction with different copper concentrations in the SRB system and SRB + Fe0 system. (a) Results of sulfate reduction ratio at different copper concentrations in the SRB system and SRB + Fe0 system. (b) Relation of sulfate reducing rates with different copper concentrations in the SRB system and SRB + Fe0 system.
increased by 559% relative to the SRB system with the copper concentration of 15 mg L − 1. It is confirmed that the supplementation of Fe 0 in the SRB culture accelerates the removal of sulfate effectively.
0
30
60
90
120
Time (minute) Fig. 4. Removal of copper with time in the SRB system, Fe0 system, and SRB + Fe0 system. (a) Change of copper with time at different copper concentrations in the SRB system and SRB + Fe0 system. (b) Comparison of copper removal ratio at copper concentration of 25 mg L− 1 in the SRB system, Fe0 system, and SRB + Fe0 system.
3.2. Copper removal
However, at the end of the experiment (120 min), the final removal ratios in the SRB system and SRB + Fe 0 system were similar; much higher than that in Fe 0 system.
3.2.1. Removal of copper In order to investigate the removal mechanism of copper, Fe 0 mentioned in Section 2.2 was added into the treatment reactor without SRB culture, which was regarded as Fe 0 system. As mentioned, during the biodegrading procedure, the sulfate and lactate were contained in the synthetic wastewater at pH 6.0 and at 36 °C. No apparent differences in the copper removal were observed in all systems (Fig. 4(a)), indicating that copper removal ratio below 0.1 mg L − 1 was achieved after 48 h at all copper concentrations in all systems. The capability and mechanism of copper removal were analyzed within 120 min at copper concentration of 25 mg L − 1. Fig. 4(b) shows that copper was removed faster in the SRB + Fe 0 system than in the SRB system and Fe 0 system. With an initial copper concentration of 25 mg L − 1 in the SRB + Fe 0 system, copper removal ratio increased from 54% to 96% after 45 min, whereas in the SRB system and Fe 0 system, copper removal ratio were only 83% and 44%.
3.2.2. Chemical composition of the precipitate In the present study, the mechanism for copper removal was apparently precipitation with sulfide. The chemical composition of the precipitate from the SRB + Fe 0 system was qualitatively analyzed with EDS at the end of the experiments. The chemical elements content of the precipitate was confirmed to be copper sulfide by the EDS spectrum shown in Fig. 5. It was revealed in Table 2 that the chemical composition of the precipitates contained large amounts of sulfur (59.81%), oxygen (28.24%), and a lesser amount of copper (5.86%), Fe, Ca, and P. In this case, the amount of sulfur was abundant to copper resulting in the complete removal of copper by means of apparent precipitation with sulfide. According to the result of XRD (data was not shown), the precipitation contains large amounts of Cu7S4, a little amount of Cu 0, CuFeS2, and Cu9Fe9S16, and insignificant amounts of oxide phases.
H. Bai et al. / International Journal of Mineral Processing 112–113 (2012) 71–76
75
621.63 534.18 1234.30 1081.84
1536.76
1453.62 1400.73
2960.27 2926.64 2856.28
80
1651.11
3294.04
Transmittance (%)
2360.53
100
Fig. 5. EDS analysis of metal precipitation sample.
In addition to precipitation with sulfide, copper might be removed through sorption on the biomass (Jalali and Balwin, 2000). In fact, SRB cells played an additional, important role in the facilitation of metal precipitation. 3.2.3. Removal mechanism The mechanisms of copper removal in the study including the effect of SRB culture and Fe 0 are rather complex. These mechanisms are integral components of biosorption, sulfide precipitation, and reductive precipitation (Gadd, 2000). The sulfate reduction ratios in the SRB system and SRB + Fe 0 system maintained at high level at low initial copper concentration indicate an efficient capability of sulfate reduction. Almost the entire copper was removed by means of precipitation with abundant sulfide produced by sulfate reduction, which was confirmed by the results of EDS analysis of the precipitate samples. It was concluded that sulfide precipitation contributes to copper removal at low initial copper concentration in the SRB system and SRB + Fe 0 system. However, the removal ratio of copper was still high with the sulfate reduction inhibited partly or completely at the high initial copper concentration in the SRB system. Copper removal to more than 95% was achieved after 216 h in the presence of 4% sulfate reduction at 25 mg L − 1 initial concentration of copper. The sulfide produced from sulfate reduction by SRB was insufficient to precipitate with copper at the high initial copper concentration. In the SRB system, it was found that the bacteria directly bind the metals in their cell walls and extracellular polymeric substances (Wang and Chen, 2006). Biosorption is either metabolism independent such as physical or chemical sorption onto the cell wall, or metabolism related such as transport and extracellular precipitation by metabolites (Gadd, 2000). Considering a high metal-reducing capability of SRB, it is suggested that SRB has a significant capacity of metal biosorption. To confirm the type of functional groups of SRB, FTIR spectra of native SRB were obtained by using a FTIR spectrophotometer. As shown in Fig. 6, The FTIR spectra of biomass indicate the presence of amino, carboxylic, hydroxyl and carbonyl groups. Display of strong broad \OH stretch carboxylic bands in the region 3294 cm − 1 and carboxylic/ phenolic stretching bands in the region of 2926 cm− 1 was observed. The peaks in the region 1651 cm− 1 were attributed to C_N, C_C and C_O stretch whereas the peaks appearing in the region 1536 cm− 1 might represent quinine O\H bond. Now the peaks appearing in the region 1356, 1081 and 1453 cm− 1 represent N\H bending, \CH3 Table 2 Element contents of sediment by EDS analysis of metal precipitation sample. Element
Composition (%)
Element
Composition (%)
O P S
28.24 1.06 59.81
Ca Fe Cu
0.78 4.25 5.86
60 4000
3500
3000
2500
2000
1500
1000
500
Wavenumber (cm-1) Fig. 6. FTIR spectra of function groups in the surface of SRB culture.
wagging, and the bending of \CH3, respectively. The biomass showed bands around 1400 and 1234 cm− 1 representing the ionized carboxylate group (COO\). The effect of Fe 0 on copper removal was studied in Fe 0 system in the absence of SRB. The copper removal was 62.2% after 120 min and reached 95% at the end of the experiment (160 h). Because standard potential of copper was more positive than iron, the reductive precipitation was predominant removal mechanism in the Fe 0 system (Li and Zhang, 2007). When a metal is converted to a lower redox state, its mobility and toxicity can be reduced. It is proposed that, for wastewater with high copper concentration, Fe 0 is an efficient reluctant for reduction of cooper. At low initial concentration of copper, the sulfate reduction ratio and copper removal were similar in the SRB system and SRB + Fe 0 system, indicating that the sulfide precipitation contributes to the copper removal. With the increasing copper concentration, sulfate reduction ratio in the SRB + Fe 0 system performed a higher level than that in the SRB system for enhanced activity of SRB by Fe 0. When the initial concentration of copper was 25 mg L − 1, the sulfate reduction ratio was still 83.46% after 160 h in the SRB + Fe 0 system. Considering the results of sulfate reduction and copper removal, it is assumed that reductive precipitation and biosorption play important roles in copper removal since the activity of SRB was constrained when copper concentration was above 25 mg L − 1. In this case, the cooperation of sulfate precipitation, biosorption and reductive precipitation occurred to remove copper in the SRB + Fe 0 system. For all copper concentrations, SRB augmented by Fe 0 was capable of removing copper completely in the SRB + Fe 0 system with sulfide precipitation and biosorption and reductive precipitation. It was demonstrated that performance of the SRB + Fe 0 system was significantly enhanced compared with the SRB and Fe 0 systems. 4. Conclusions The activity of SRB enhanced by Fe 0 played an important role in copper removal in the presence of high concentrations of copper (20 mg L − 1 and 25 mg L − 1). The 84.36% of sulfate reduction ratio in SRB + Fe 0 system was much higher than the 5.6% in the SRB system after 216 h at 25 mg L − 1of copper. The sulfate reduction rate at 82 mg L − 1 h − 1 was achieved in the presence of 25 mg L − 1 of copper in SRB + Fe 0 system, whereas only 5.6 mg L − 1 h − 1 of sulfate reduction rate was observed in the SRB system. Similar capability of copper removal was observed in all systems, and the overall copper removal was around 99.5% after 40 h in all three systems, whereas the SRB + Fe 0 system exhibited a better performance of copper removal than the other systems within 120 min. In the copper removal experiment, EDS analysis indicated the
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formation of a dominating crystalline structure that was mainly composed of Cu7S4. Sulfide precipitation and biosorption contribute to the copper removal in the SRB system. In contrast, reductive precipitation played an important role in the removal of copper in the Fe 0 system. The removal of copper was most effective in the SRB + Fe 0 system due to the cooperation of sulfide precipitation, biosorption and reductive precipitation. Performance of the SRB + Fe 0 system was significantly enhanced compared with the SRB and Fe 0 systems. Acknowledgments The authors are grateful for the financial support of National Nature Science Foundation of China (Project Nos. 21077075 and 20106014). References Akcila, A., Koldasb, S., 2006. Acid Mine Drainage (AMD): causes, treatment and case studies. J. Clean. Prod. 14, 1139–1145. Bayrakdar, A., Sahinkaya, E., 2009. Performance of sulfidogenic anaerobic baffled reactor (ABR) treating acidic and zinc-containing wastewater. Bioresour. Technol. 100, 4354–4360. Viggi, C., Pagnanelli, F., Cibati, A., Uccelletti, D., Palleschi, C., Toro, L., 2010. Biotreatment and bioassessment of heavy metal removal by sulphate reducing bacteria in fixed bed reactors. Water Res. 44, 151–158. Teclua, D., Tivchev, G., Laing, M., Wallis, M., 2008. Bioremoval of arsenic species from contaminated waters by sulphate reducing bacteria. Water Res. 42, 4885–4893. Sahinkaya, E., Gunes, F.M., Ucar, D., Kaksonen, A.H., 2011. Sulfidogenic fluidized bed treatment of real acid mine drainage water. Bioresour. Technol. 102, 683–689. Sahinkaya, E., Gungor, M., 2010. Comparison of sulfidogenic up-flow and down-flow fluidized-bed reactors for the biotreatment of acidic metal-containing wastewater. Bioresour. Technol. 101, 9508–9514. Pagnanelli, F., Viggi, C.C., Toro, L., 2010. Isolation and quantification of cadmium removal mechanisms in batch reactors inoculated by sulphate reducing bacteria: biosorption versus bioprecipitation. Bioresour. Technol. 101, 2981–2987. Cabrera, G., Pérez, R., Gómez, J.M., Ábalos, A., Cantero, D., 2006. Toxic effects of dissolved heavy metals on Desulfovibrio vulgaris and Desulfovibrio sp. strains. J. Hazard. Mater. 135, 40–46. Gadd, G.M., 2000. Bioremedial potential of microbial mechanisms of metal mobilization and immobilization. Curr. Opin. Biotechnol. 11, 271–279.
Wang, J., Chen, C., 2006. Biosorption of heavy metals by Saccharomyces cerevisiae: a review. Biotechnol. Adv. 24, 427–451. Jalali, K., Balwin, S.A., 2000. The role of sulfate-reducing bacteria in copper removal from aqueous sulfate solutions. Water Res. 34, 797–806. Malaeb, L., Ayoub, G.M., 2011. Reverse osmosis technology for water treatment: state of the art review. Desalination 267, 1–8. Zhang, L.H., De Schryver, P., De Gussemea, B., De Muynck, W., Boon, N., Verstraete, W., 2008. Chemical and biological technologies for hydrogen sulfide emission control in sewer systems: a review. Water Res. 42, 1–12. Martins, M., Faleiro, M.L., Barros, R.J., Raquel Veríssimo, A., Barreiros, M.A., Clara Costa, M., 2009. Characterization and activity studies of highly heavy metal resistant sulphate-reducing bacteria to be used in acid mine drainage decontamination. J. Hazard. Mater. 166, 706–713. Matlock, M., Howerton, B.S., Atwood, D.A., 2002. Chemical precipitation of heavy metals from acid mine drainage. Water Res. 36, 4757–4764. Alexandrinoa, M., Macíasb, F., Costaa, R., Gomesc, N.C.M., Canárioa, A.V.M., Costa, M.C., 2011. A bacterial consortium isolated from an Icelandic fumarole displays exceptionally high levels of sulfate reduction and metals resistance. J. Hazard. Mater. 137, 162–370. Lindsay, M.B.J., Ptacek, C.J., Blowes, D.W., Douglas Gould, W., 2008. Zero-valent iron and organic carbon mixtures for remediation of acid mine drainage: batch experiments. Appl. Geochem. 23, 2214–2225. Wilkin, R.T., McNeil, M.S., 2003. Laboratory evaluation of zero-valent iron to treat water impacted by acid mine drainage. Chemosphere 53, 715–725. Curteanu, S., Piuleac, C.G., Godini, K., Azaryan, G., 2011. Modeling of electrolysis process in wastewater treatment using different types of neural networks. Chem. Eng. J. 172, 267–276. Karri, S., Alvarez, R.S., Field, J.A., 2005. Zero-valent iron as an electron donor for methanogenesis and sulfate reduction in anaerobic sludge. Bioresour. Technol. 92, 810–819. Cochran, W.G., 1950. Estimation of bacterial densities by means of the most probable number. Biometrics 6, 105–116. Ngah, W.S.W., Hanafiah, M.A.K.M., 2008. Removal of heavy metal ions from wastewater by chemically modified plant wastes as adsorbents: a review. Bioresour. Technol. 99, 3935–3948. Li, X., Zhang, W., 2007. Sequestration of metal cations with zerovalent iron nanoparticles — a study with high resolution X-ray photoelectron spectroscopy (HR-XPS). J. Phys. Chem. C 111, 6939–6946. Yao, X., Kang, Y., Lee, D., Feng, Y., 2008. Bioaugmented sulfate reduction using enriched anaerobic microflora in the presence of zero valent iron. Chemosphere 7, 1436–1441.