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Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor
Accepted Manuscript Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor Siyuan Zhai, Yinxin Zhao, Min Ji, Wenfang Qi PII: DOI: Reference:
S0960-8524(17)30148-7 http://dx.doi.org/10.1016/j.biortech.2017.01.076 BITE 17604
To appear in:
Bioresource Technology
Received Date: Revised Date: Accepted Date:
2 November 2016 19 January 2017 22 January 2017
Please cite this article as: Zhai, S., Zhao, Y., Ji, M., Qi, W., Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor, Bioresource Technology (2017), doi: http://dx.doi.org/10.1016/ j.biortech.2017.01.076
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Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor Siyuan ZHAI a, Yinxin ZHAO a,b*, Min JI a,b, Wenfang QI a a
School of Environmental Science and Engineering, Tianjin University, Tianjin
300350, China b
Tianjin Engineering Center of Urban River Eco-Purification Technology, Tianjin
300350, China Abstract: A spiral fiber based biofilm reactor was developed to remove nitrate and chromate simultaneously. The denitrification and Cr(VI) removal efficiency was evaluated with synthetic groundwater (NO3--N=50 mg/L) under different Cr(VI) concentrations (0~1.0 mg/L), carbon nitrogen ratios (C/N) (0.8~1.2), hydraulic retention times (HRT) (2~16 h) and initial pHs (4~10). Nitrate and Cr(VI) were completely removed without nitrite accumulation when the Cr(VI) concentration was lower than 0.4 mg/L. As Cr(VI) up to 1.0 mg/L, the system was obviously inhibited, but it recovered rapidly within 6 days due to the strong adaption and domestication of microorganisms in the biofilm reactor. The results demonstrated that high removal efficiency of nitrate (≥ 99%) and Cr(VI) (≥ 95%) were achieved at lower C/N = 0.9, HRT = 8 h, initial pH = 7, and Cr(VI) = 1.0 mg/L. The technology proposed in present study can be alternative for simultaneous removal of co-contaminants in groundwater. Key Words: Denitrification, Cr(VI) removal, Biofilm reactor, Groundwater *
Corresponding author: Yingxin Zhao. E-mail: [email protected] (Y.Zhao); Tel/Fax: +86 22 2740 6057
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Introduction Groundwater is one of the main water resources, which is widely used as drinking water in most countries of the world. However, in recent years, the co-existence of different oxidized contaminants such as nitrate, sulfate, pesticide and chromate in natural water or wastewater becomes a common and growing problem (Li et al., 2016). In the nature, no matter the riverine system (Olivares et al., 2010), soil system (Katarzyna et al., 2016; Liu et al., 2015) and groundwater system (Babiker et al., 2004), the long-term combined contamination were more harmful to the environment and the toxicity of compound pollution could change the microbial community structure. Nitrate is the most ubiquitous chemical contaminant in groundwater due to excessive utilization of nitrogenous fertilizer in agriculture and discharge of domestic and industrial waste (Almasri et al., 2004; Wang et al., 2009; Chen et al., 2013).The presence of nitrate in groundwater is increasing worldwide, because it is not easily removed by conventional drinking water treatment (Glass et al., 1998). Nitrate can cause acute health problems when it is reduced into nitrite, which in turn can react with secondary or tertiary amines to produce carcinogenic nitrosamines. The US Environmental Protection Agency's standard has set the maximum contaminant level of nitrate at 10 mg/L and the same values are also proposed by China (Standards for drinking water quality, GB5749-2006).The conventional physicochemical methods used to eliminate nitrate from water are ion exchange (Bae et al., 2002), reverse osmosis (Schoeman et al., 2003) and electro-dialysis (Peel et al., 2003). In regards to 2
biological methods, biological denitrification is widely applied for nitrate removal from groundwater attributing to its high efficiency and low cost. Chromium, another environmental pollutant of interest in groundwater reservoirs, is one of the most abundant and toxic metals that cause the pollution of groundwater and soil due to its frequent industrial application, such as wood preservation, electroplating, alloy production and leather tanning (Sahinkaya et al., 2013). In the aquatic environment, chromium primarily exists in trivalent Cr(III) and hexavalent forms Cr(VI). Cr (VI) is highly soluble and mobile, which shows acute toxicity, mutagenicity and carcinogenicity in ecosystem (Somasundaram et al., 2009). While the relation between Cr(III) and Cr(VI) depends strongly on pH and the oxidative properties of the location. As a result, Cr(VI) reduction and subsequently immobilization of the generated Cr(III) is an effective method to remove and detoxify Cr(VI) (Li et al., 2016). The treatment of Cr(VI) polluted wastewater including chemical precipitation, electrochemical treatment, reverse osmosis and ion exchange (Barrera et al., 2012). However, the aforementioned methods present disadvantages such as high cost, energy consumption or production of chemical sludge, which makes them not suitable for removing low concentration of Cr(VI) in groundwater (Ganguli et al., 2002). Hence, biotransformation of Cr(VI) to the less toxic Cr(III) may be an alternative promising approach for groundwater treatment. Biological reduction of different oxidized contaminants to harmless or immobile forms is considered as a promising approach to solve compound pollution problems. Recently, Cr(VI) reduction by bacteria was found to have indivisible relationships 3
with the physiological electron acceptor (oxygen or nitrate) consumption (Somasundaram et al., 2009; He et al., 2015). In addition, some of previous studies have reported the simultaneous reduction of nitrate and chromate (Li et al. 2016; Chung et al., 2006; Miao et al., 2015; Sahinkaya et al., 2014). Chung et al. (2006) proposed that the simultaneous removal of nitrate and chromate in drinking water could be achieved using H2 as an electron donor in a H2-based membrane biofilm reactor. Sahinkaya et al. (2014) accomplished the removal of nitrate and Cr(VI)-containing wastewater in sulfur-based autotrophic denitrifying column bioreactors. Complete autotrophic denitrification was obtained at a nitrate loading rate of 0.1 g NO3--N/(L·d) and the influent Cr(VI) concentration below 0.5 mg/L. Lower removal rates of nitrate and chromate were attained, while the problem of secondary pollution by sulfur-based autotrophic denitrifying couldn’t be ignored. Recently, a solid substrate as an alternative to the soluble carbon source was developed, which could not only provide constantly reducing power for reduction of oxidized pollutants when attacked by microbial enzyme but also serve as support for biofilm formation and development. From the finding of Li, an up-flow packed-bed bioreactor was constructed to investigate the simultaneous removal of chromate. In this system, a biodegradable meal box was taken as carbon source and biofilm carriers, and complete denitrification and Cr(VI) reduction could be achieved with high Cr(VI) concentrations (2.0~50.0 mg/L) and longer HRT (>17h) (Li et al. 2016). However, continuous and stable release of organic carbon was also a problem by taking biodegradable meal box as solid carbon source. Therefore, more efforts should be 4
dedicated to develop a simple and efficient biofilm reactor to overcome the main disadvantages of existing technology, such as the high cost, low removal efficiency, secondary pollution and the complexity of reactor operation.. In this study, a spiral fiber carrier based biofilm reactor was developed to remove nitrate and chromate simultaneously. The simultaneous removal efficiencies of nitrate and chromate under different Cr(VI) concentrations, carbon and nitrate ratios (C/Ns), hydraulic retention times (HRTs) and initial solution pHs were investigated. In addition, the main Cr speciation and distribution around the biofilm were also analyzed. 2. Materials and methods 2.1 Experimental apparatus Fig.1 shows a set of lab-scale biofilm reactor used in this study. The main part of the reactor was an organic glass cylinders (diameter 300 mm, height 240 mm) with an effective volume of 7.5 L. The main reactor compartments consisted of a water collector (diameter 50 mm, height 200 mm), a water carrier and a water bath outer for heat preservation. The water collector was installed in the center of the cylinder, 108 holes (diameter 5 mm, distance between hole centers 20 mm) were drilled symmetrically in the collector wall. Each hole was filled with the carriers and the carriers bound on the collector. Peristaltic pumps were used to control the flow rate of influent and effluent. Synthetic groundwater was pumped from the influent tank into the biofilm reactor through the bottom inlet of the reactor. Influent flowed through the biofilm and effluent was drawn from the water collector. While the biofilm also 5
worked as a filter preventing suspended solids (SS) and other impurities into the treated water, therefore it could save a sedimentation tank or membrane filtration. 2.2 Chemicals and materials Synthetic contaminated groundwater (per liter of tap water) contained NaNO3, phosphate buffer solution and potassium dichromate. CH3OH was added as required carbon source to maintain the desired C/N. The original pH of the influent solution was normally about 7.0±0.2. All chemical reagents used were analytical grade without further purification. All solid reagents were weighed using analytical balance (Sartorius BS 124S, Germany) and liquid reagents were measured via pipettes with specific ranges. A mixed culture was originally collected from anaerobic sludge, which was obtained from the secondary sedimentation tank of Zhang-Gui-Zhuang Sewage Treatment Plant (Tianjin, China). In recent study, the anaerobic sludge collected from the same wastewater treatment plant contains some denitrifying bacteria which was proved by the results of the high-throughput sequencing. The carrier provided by Shijiazhuang green environmental protection technology co., Ltd. is composed of cotton, vinylon, polyvinylidene chloride and polypropylene fiber, and its shape is imitation of spiral aquatic plants type. The characteristics of the spiral fiber based biofilm reactor are strong adhesion of microorganisms, low cost and easy management. 2.3 Experiment procedure The biofilm reactor was set up to achieve the simultaneous removal of nitrate and 6
chromate under different operating conditions. The biofilm reactor was inoculated with anaerobic sludge and operated at 25-28 oC during the experiments. Biofilm formed gradually and a dark gray color was observed on the carrier within 10 days. During the whole operating periods, NO3--N concentration of influent was 50 mg/L. Then the biofilm reactor began to operate as indicated in Table 1. The biofilm reactor was operated for around 126 days under 4 different effect factors. In the first period, a low Cr(VI) concentration was kept at 0.1 mg/L for 12 days to enrich bacterial growth and promote biofilm formation. Then the influence of Cr(VI) concentrations (0.1, 0.2, 0.3, 0.4, 0.5 and 1.0 mg/L) on simultaneous nitrate and Cr(VI) removal was investigated. In phases 2, 3 and 4, the influent Cr(VI) concentration was determined at 1.0 mg/L. To study the effects of C/N and HRT on denitrification and Cr(VI) reduction, C/N was gradually decreased from 1.2 to 0.8, and the HRT was controlled at 16, 12, 8, 4, and 2 h. In the last period, the initial pHs of the system were conducted at 4, 7 and 10. During these procedures, the operating conditions of simultaneous nitrate and Cr(VI) removal were optimized. 2.4 Analysis method The influent and effluent water samples’ indicators of each cycle reaction were detected, including pH, CODCr, Cr(VI), Cr(III), NO3--N, NH4+-N and NO2--N. The pH in the influent and effluent was determined by a pH meter (Inesa PHSJ-3F, China). The chemical oxygen demand (COD) was measured using the dichromate method with HACH DR2800 (Rice et al., 2013). Cr(VI), NO3--N, NH4+-N and NO2--N were determined in accordance with standard methods. 7
A thermometer was set into the reactor to monitor the solution temperature during the experiments. 3. Results and discussion 3.1 Denitrification performance under different concentrations of Cr(VI) stress High Cr(VI) concentration in wastewater would conducted toxicity on activated sludge process, and the system was obviously affected as it was above 5 mg/L (Vaiopoulou et al., 2012). Comparing to wastewater, relatively lower Cr(VI) concentrations exist in groundwater environment, normally lower than 1.0 mg/L. Previous studies reported that 0.5 mg/L of Cr (VI) could impact on denitrification performance (Li et al. 2016). In the present study, different Cr(VI) concentrations (0.1, 0.2, 0.3, 0.4, 0.5 and 1.0 mg/L) were fed into the biofilm reactor during the first stage (1-38 d) to investigate the impact resistance of the system. The concentrations of NO3--N, NO2--N, NH4+-N and Cr(VI) in effluent under different influent Cr(VI) conditions were shown in Fig. 2. Denitrification performance under Cr(VI) stress was presented in Fig.2 (a). Almost complete denitrification was achieved when the concentrations of influent Cr(VI) were increased from 0.1 to 0.4 mg/L, which indicated that low concentrations of Cr(VI) had no obviously negative effects on denitrification. Increasing feed Cr(VI) concentration to 0.5 mg/L, the effluent nitrate concentrations increased to 1.39 mg/L, significantly responding the inhibiting effects of higher Cr(VI) on denitrification of the system. However, the nitrate removal efficiency recovered in two days and the nitrite concentration still remained at very low levels (≤ 0.075 mg/L). Further 8
increasing influent Cr(VI) concentration from 0.5 mg/L to 1.0 mg/L, obvious inhibiting effects were observed on denitrification. The nitrate removal rate decreased from almost 100% to 89.44%, and 0.78 mg/L NO2--N accumulated at the same time. The significant lag and inhibition of nitrate reduction herein were ascribed to the electron competition between nitrate and Cr(VI) (He et al., 2015). Another was that the Cr(VI) reduction process gave rise to reactive free radicals, such as Cr(V) and O2·, which further caused oxidative damage on DNA, RNA and proteins of the bacterial cells and severely influenced the enzyme synthesis and bacterial metabolism (Valko et al., 2005). After a period (about 7 days) operation, the microorganisms of the system may gradually produce a certain degree tolerance on higher Cr(VI) load through adaptation and domestication. Finally, the restoration of nitrate removal efficiency gradually improved and the effluent NO3--N concentration was below 0.5 mg/L. Bacteria reduce nitrate into nitrite with nitrate reductase enzymes (Nap or Nar), then nitrite can be transformed into ammonia through dissimilatory nitrate reduction to ammonia (DNRA) by nitrite reductase enzymes (Stolz et al., 2002). Therefore, the NH4 +-N variation (Fig. 2 (a)) was also analyzed during the whole operating periods of different Cr(VI) concentrations. At relatively low initial Cr(VI) level (0.1 mg/L and 0.2 mg/L), NH4+-N of the system accumulated to 2.53 mg/L. Then, the effluent ammonia concentrations were kept below 1.0 mg/L with increasing Cr(VI) concentration. The cytochromes of multiheme nitrite reductase (Nrf) can be readily oxidized by Cr(VI) thus rendering them inoperable (Stolz et al., 2011). As a result, the activity of the enzymes involved in DNRA will be suppressed by Cr(VI) adding. 9
The variations of influent and effluent Cr(VI) were plotted in Fig. 2(b). The effluent Cr(VI) rose obviously with increasing Cr (VI) load, but the system recovered after several cycles. During the reactor operation, the effluent Cr(VI) concentrations (≤ 0.05 mg/L) were even lower than the drinking water standard when the influent Cr(VI) concentration steadily increased up to 0.5 mg/L. As the influent Cr(VI) concentration was increased further, both denitrification and Cr(VI) removal capacity of the reactor decreased and effluent Cr(VI) reached around 0.18 mg/L at Cr(VI) concentration of 1.0 mg/L. Although a fluctuation of Cr(VI) decline occurred at each changed Cr(VI) load, Cr(VI) concentration finally reached below 0.05 mg/L based on the good adaption ability of the reactor. Cr(VI) is more stable under oxidizing conditions and Cr(III) is more stable under reducing conditions. The implementation of microbial chromium bioremediation has encountered, and anaerobic reduction of Cr(VI) can be mediated by several different microorganism. Peter (2012) reported three ways of biotransformation of nitrate and chromate contaminated environments: (1) competitive alternative electron acceptor; (2) co-metabolism; (3) induction of specific proteins and pathways involved in oxidation/reduction reactions. For the first way, the Cr(VI) reductase (ChrR) has more competition capability for electrons than nitrate reductase (Salamanca et al., 2013; Kourtev et al., 2009). In Fig.3 (a), effluent average CODCr concentration showed a significantly rise from 21.42 mg/L to 39.59 mg/L when the influent Cr(VI) was from 0.1 mg/L up to 0.2 mg/L. The decline of methanol utilization may be due to the inhibition of denitrification activity by Cr(VI) stress, and subsequently the amounts of dissolved 10
organic carbon (DOC) released exceeded what denitrification needed (Bae et al., 2002). However，it was worth noting that the effluent COD concentrations decreased with further increasing influent Cr(VI) concentration from 0.2 mg/L to 1.0 mg/L after a period operation. It proved that the biofilm reactor could tolerate higher chromium concentrations under heterotrophic conditions. The influent and effluent pH variation of the reactor were presented in Fig. 3 (b). During the overall operating periods under different Cr(VI) concentrations, no obvious effects of increasing influent Cr(VI) loading on effluent pH were observed. The initial pH maintained 7±0.2, the effluent pH was slightly above 7.5 due to the alkalinity generation of denitrification process (Equation 1). 6NO3-+5CH3OH→3N2+7H2O+5CO2+6OH-
(1)
3.2 Effect of C/N ratio on simultaneous nitrate and Cr(VI) removal
In this reactor, methanol was supplemented as carbon source. Through changing the methanol dosage, different C/N (1.2, 1.0, 0.9 and 0.8 mg/L) were studied in the biofilm reactor during the second stage (39~63 d) to investigate the impact resistance of the system. The concentrations of NO3--N, NO2--N and Cr(VI) in effluent under different C/N conditions were shown in Fig. 4. As seen in Fig.4(a), the effluent nitrate concentrations were quite low and almost complete denitrification was attained at relative higher C/N operation parameter (1.2~0.9). When C/N was above 0.9, the nitrite concentration in effluent was always below detection limit (0.005 mg/L NO2—N). Between days 54 and 59 (C/N = 0.8), nitrate and nitrite concentrations increased to 5.58 mg/L and 4.84 mg/L in the effluent, 11
respectively. This decline of denitrification efficiency may be due to limiting amount of methanol in the influent. At the same time, the Cr(VI) reduction process also responses to the shortage of the carbon source, and removal rate decreased to 80%. Sahinkaya did similar research about simultaneous bioreduction of nitrate and chromate under mixotrophic denitrification conditions (Sahinkaya et al., 2013). In their study, the optimal C/N ratio was 1.33 with combined heterotrophic and sulfur based autotrophic processes, and almost complete denitrification was achieved at influent NO3−–N and Cr(VI) concentrations of 75 mg/L and 10 mg/L, respectively. In comparison with Sahinkaya’s study, the biofilm reactor used in the present study performed better with lower C/N ratio of 0.9, indicating a cost advantage for its application. In order to recover denitrification and Cr(VI) reduction capabilities of the system, C/N was increased to 0.9 on day 59 of operation to create the optimum conditions for the microorganisms, thereby ensuring a stable and efficient operation of the biofilm reactor. Under the optimum operating condition, the denitrification and Cr(VI) reduction performance of the system was recovered in 3 days. 3.3 Effect of HRT on simultaneous nitrate and Cr(VI) removal
The impact of the key operating parameter-HRT on the nitrate and Cr(VI) removal performance of biofilm reactor was also investigated. With the presence of 50 mg/L nitrate, 210 mg/L COD and 1.0 mg/L Cr(VI) in the influent, the removal efficiencies of nitrate and COD at varying HRTs were shown in Fig.5. The average concentration of Cr(VI) and Cr(III) in effluent can be seen in Tab. 2. 12
As HRT decreased from 16 to 8 h, the removal efficiencies of nitrate, COD and Cr(VI) were in the ranges of 97~100%, 82.2~95.6% and 88~100%, respectively. There were no significant differences of pollutants removal efficiency as the HRT= 16, 12, 8 h, so the hydraulic retention time of 8 h could be identified as a suitable reaction parameters. Meanwhile, no accumulation of nitrite was detected, which indicated that the system was stable. The biological Cr(VI) reduction approach was controversial process for different researchers of different systems. The reduction pathway of Cr(VI) maybe reduced the biological toxicity to a certain extent, but not accomplish the thorough removal of Cr(VI) (Singh et al., 2011). In some studies, generated Cr(III) by Cr(VI) reduction were achieved almost 100% immobilization rate by biofilm reactor (Pan et al. 2014). While, Novotnik (2014) reported that Cr(VI) reduction mainly occurred in the intercellular portion. In our system, the Fig. 5(c) was shown that :one part of hexavalent chromium were removed completely; the other part of Cr(VI) transformed to Cr(III) and the Cr(III) concentrations were accumulated at the range of 0.02~0.21 mg/L when HRTs were higher than 8 h. Further decrease of HRT to 4 and 2 h, concomitantly, the contaminants removal rates were significantly decreased. Particularly from day 100 to 110, further decreasing HRT to 2 h led to a remarkable increase of nitrate (26.61 mg/L) and nitrite (9.18 mg/L) in the effluent and the denitrification rates decreased sharply to 60% (Fig.5(a) and Tab.2). While the COD removal rate collapsed rapidly from 90% to 70%. Besides, decreasing HRT led to an obvious decline of Cr(VI) removal rate to 48% 13
(Tab.2) and effluent total Cr = 0.82 mg/L (supported in the Fig.5c) was almost equal to influent Cr(VI) amount with little Cr(VI) transformation to Cr(III). To remove NO3--N and Cr(VI) effectively, the HRT was controlled at 8 h in the following studies. 3.4 Effect of initial pH on simultaneous nitrate and Cr(VI) removal
In order to recover denitrification and Cr(VI) reduction capability of the system and keep stabilization of the biofilm reactor for the next impact factor investigating, the HRT increased to 8 h and operating for 5 days. The pH of medium also played an important role in nitrate and Cr(VI) reduction. As shown in Fig.6, the system could effectively remove both nitrate and Cr(VI) at pH 7 and 10, which more than 95% of Cr(VI) removal efficiency was achieved and 99% nitrate removal efficiency was reached. The Cr(III) precipitation produced by Cr(VI) transformation was at a very low level (≤ 0.015 mg/L) during the operating periods under different initial pHs. Similarly, He et.al indicated that both Cr(VI) and nitrate could be removed effectively at pH from 8 to 9, in which more than 80% of Cr(VI) removal was achieved and nitrate was depleted (He et al., 2015). In addition, effluent pHs were steadily in the range of 7.5 to 8.0. As the initial pH was conducted to 4, the acidic condition distinctly defeated the nitrate and Cr(VI) reduction ability. While the NO3--N concentration reached 1.76 mg/L and Cr(VI) accumulated to 0.15 mg/L. The growth and activities of microorganism would be suppressed significantly at the condition of strong alkaline (pH > 10) or acidic (pH < 6) which distinctly defeated the Cr(VI) and nitrate reduction ability (He et al., 2015). 14
Generally, the pH may be a factor affecting another pathway DNRA. Some previous investigations reported that the optimum pH was 6.5 and 7.5 for NO2--N and NO3--N reduction, and the higher DNRA was generally observed in alkaline environments (Stevens et al. 1998; Rütting et al. 2011; Zhang et al. 2015).With the system changing from acidic to alkaline condition in our system, the obviously increasing NH4+-H concentration was found by the DNRA pathway. This phenomenon was consistent with the findings of Zhang (2015), who stated that the DNRA process was negligible under acidic condition(pH 4.7), but was mor prevalent in neutral (pH 6.2) and alkaline (pH 8.2) environments. 3.5 Comparison of nitrate and Cr(VI) removal with different systems
Tab.3 presented a comparison between the reactor used in this study and other system used for the simultaneous removal of nitrate and Cr(VI), including the information about the main operational parameters and removal efficiencies of each system. It can be seen from Tab.3 that the biofilm reactor in the present study has significant advantages. The obvious superiority lies in high removal efficiency of contaminants (≥ 99% nitrate and ≥ 95% Cr(VI)), lower carbon and nitrogen ratio (0.9) and shorter hydraulic retention time (8 h). The operational costs of this system are mainly related to carbon source, therefore the use of a low carbon to nitrogen ratio reduces the operational costs for its application in the treatment of co-contaminants present in groundwater. It is apparent that the biofilm reactor is favorable for treatment of nitrate and Cr(VI) co-contaminated groundwater. 15
4. Conclusions The simultaneous removal of nitrate and Cr(VI) was achieved by a spiral fiber based biofilm reactor. Low Cr(VI) concentration (≤0.4 mg/L) had no obviously negative effects on nitrate removal. The denitrification performance was inhibited as the Cr(VI) concentration increased to 1.0 mg/L, but the restorations of nitrate and Cr(VI) removal efficiency gradually improved more than 99% and 95% after a period operation. The optimum C/N, HRT and initial pH were observed at 0.9, 8 h and pH=7. It indicated that the method had promising application for treatment of nitrate and Cr(VI) co-contaminated groundwater.
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Acknowledgments： ： Authors are grateful to the financial support from the National Key Technology Research and Development Program (Project No. 2015BAL04B03, China). and Critical Patented Projects in the Control and Management of the National Polluted Water Bodies (Project No. 2015ZX07203-011, China)
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area in South Africa. Desalination, 155(1), 15-26. Singh, S. K., Bansal, A., Jha, M. K., Dey, A., 2011. An integrated approach to remove Cr(VI) using immobilized Chlorella minutissima grown in nutrient rich sewage wastewater. Bioresour. Technol., 104(1), 257–265. Somasundaram, V., Philip, L., Bhallamudi, S. M.. 2009. Experimental and mathematical modeling studies on Cr(VI) reduction by CRB, SRB and IRB, individually and in combination. J. Hazard. Mater., 172(2-3), 606–617. Stevens, R. J., Laughlin, R. J., Malone, J. P., 1998. Soil pH affects the processes reducing nitrate to nitrous oxide and di-nitrogen. Soil. Bio. Biochem., 30, 1119-1126 Stolz, J. F., Ass, P. B., 2002. Evolution of nitrate reductase: molecular and structural variations on a common function. Chembiochem, 3(2-3), 198-206. Stolz, J. F., 2011. Nitrate enhanced microbial Cr(VI) reduction-final report. Office of Scientific & Technical Information Technical Reports. Vaiopoulou, E., Gikas, P., 2012. Effects of chromium on activated sludge and on the performance of wastewater treatment plants: A review. Water Res., 46(3), 549-570. Valko, M., Morris, H., Cronin, M. T., 2005. Metals, toxicity and oxidative stress. Curr. Med. Chem., 12(10), 1161-208. Wang, Q., Feng, C., Zhao, Y., Hao, C., 2009. Denitrification of nitrate contaminated groundwater with a fiber-based biofilm reactor. Bioresour. Technol., 100(7), 2223–2227. Zhang, J., Lan, T., Müller, C., Cai, Z., 2015. Dissimilatory nitrate reduction to ammonium (DNRA) plays an important role in soil nitrogen conservation in neutral and alkaline but not acidic rice soil. J. Soil. Sediment., 15(3), 523-531. 21
Figure Captions Fig.1. Experimental apparatus Fig.2. Denitrification and Cr(VI) removal under different concentrations of Cr(VI) Fig.3. Variations in effluent CODCr concentrations and pH under different concentrations of Cr(VI) Fig.4. Denitrification and Cr(VI) removal under different C/N Fig.5. The biofilm reactor removal efficiency under different HRTs Fig.6.The impact of initial pH on system denitrification and Cr(VI) reduction performance
22
6 11
b 7 9
a
3
5
10
4 2 1
12
8
Fig.1 Experimental apparatus (1.reactor; 2. water bath outer; 3.water collector; 4.carriers; 5.wastewater inlet; 6.treated water outlet; 7.overflow pipe; 8. constant temperature circulating water inlet; 9. constant temperature circulating water outlet; 10,11. peristaltic pump; 12. influent tank; a. influent pipe; b. effluent pipe.)
23
6
-
NO3 -N
5 N concentration / (mg/L)
1.0
Influent Cr(VI) -
+
NO2 -N
NH4 -N
4
0.5
3 2
0.0
1 0
0
10
20
30
Influent Cr(VI) /(mg/L)
(a)
-0.5 40
Time/(d)
0.30
Influent Cr(VI)
Influent Cr(VI)/(mg/L)
Effluent Cr(VI)
0.25
0.9 0.20 0.15
0.6
0.10 0.3 0.05 0.0
0
10
20
30
Effluent Cr(VI)/(mg/L)
(b)1.2
0.00 40
Time/(d)
Fig.2 Denitrification and Cr(VI) removal under different concentrations of Cr(VI)
24
(a)
80
Influent COD Cr=205~215 mg/L
Effluent CODCr / (mg/L)
Effluent COD Cr
60
40
20
0
0.1
0.2
0.3
0.4
0.5
1.0 6
Influent Cr(VI)/(mg/L)
1.0
Influent Cr(VI) / (mg/L)
9.0
Influent Cr(VI) pH variation
8.5
0.8
8.0
0.6
7.5
Initial pH=7±0.2
0.4
7.0
0.2 0.0
pH variation
(b) 1.2
6.5
0
10
20
30
6.0 40
Time/(d)
Fig.3 Variations in effluent CODCr concentrations and pH under different concentrations of Cr(VI)
25
(a)
8
1.4
C/N ratio NO2--N
6
1.2 1.2
4
1.0 0.9
1.0
0.8
2
0
0.9
40
44
48
52
56
0.8
60
64
C/N variation
N concentration /(mg/L)
NO3--N
0.6
Time/(d)
(b)0.25
1.4
C/N ratio Effluent Cr(VI)
1.2 1.2
0.15 0.10
1.0
0.9
1.0
0.9 0.8
0.8
0.05 0.00
40
44
48
52
56
60
64
C/N variation
Effluent Cr(VI) /(mg/L)
0.20
0.6
Time/(d)
Fig.4 Denitrification and Cr(VI) removal under different C/Ns
26
N concentration / (mg/L)
25
HRT NO3 -N
20 + NH4 -N
NO2 -N
16
20
12
15 8
10
4
5 0
70
80
90
100
110
HRT variation /(h)
(a) 30
0
Time/(d)
75
20
HRT Effluent CODCr
16
60
12
45
8
30
30mg/L 15 0 60
70
80
90
100
110
4
HRT variation / (h)
CODCr concentration / (mg/L)
(b) 90
0
Time / (d)
Cr(VI) Cr concentration / (mg/L)
20
HRT Cr(III)
0.8
15
0.6 10 0.4 5
0.2 0.0
70
80
90
100
110
HRT variation / (h)
(c) 1.0
0
Time / (d)
Fig.5 The biofilm reactor removal efficiency under different HRTs
27
contaminant concentrations / (mg/L)
4
NO 3 - -N
NO 2 - -N
NH 4 + -N
Cr(VI)
3
2
1
0
41
2
7
initial pH
3
10
Fig.6 The impact of initial pH on system denitrification and Cr(VI) removal performance
28
Table Captions Tab.1. Operating periods of biofilm reactor Tab.2. Nitrate and Cr(VI) reduction activity of different HRTs Tab.3. Simultaneous removal of nitrate and chromate in different reaction system
29
Tab.1 Operating periods of biofilm reactor Periods Operating dates/(d) NO3--N/(mg/L) Cr(VI)/(mg/L) C/N HRT/(h) Initial pH
1
2
3
4
1-38 50 0.1,0.2,0.3,0.4,0.5,1.0 1.2 8 7
39-63 50 1 0.8,0.9.1.0,1.2 8 7
64-110 50 1 0.9 2,4,8,12,16 7
115-136 50 1 0.9 8 4,7,10
30
Tab.2 Nitrate and Cr(VI) reduction activity of different HRTs HRT(h) Nitrate removal rates (%)
16
12
8
4
2
100
100
99
94
60
Effluent Cr(VI) (mg/L)
0.036±0.02 0.041±0.01 0.048±0.05
0.32±0.12 0.52±0.24
Effluent Cr(III) (mg/L)
0.009±0.01 0.012±0.04 0.008±0.07
0.18±0.10 0.24±0.08
31
Tab.3 Simultaneous removal of nitrate and chromate in different reaction system
Reaction system Biofilm reactor Expanded granular sludge bed reactor Up-flow packed-bed bioreactor Autotrophic column bioreactor Heterotrophic column bioreator
NO3--N (mg/L)
Cr(VI) (mg/L)
50
1.0
200
10
100
2.0
75
2.0
75
2.0
Operating conditions (Methnol)C/N=0.9, HRT=8 h Liquid up-flow velocity =2.0 m/h (BMB)COD=400 mg/L HRT=17 h, (Methnol)C/N=2.85, HRT=12h (Methnol)C/N=3.41, HRT=12h
32
Nitrate/Cr(VI) removal rate(%) ≥99/95 ≥99.5/95 ≥99/95 ≥64/≥99/-
Reference This study Miao et al., 2015 Li et al., 2016 Sahinkaya et al., 2014 Sahinkaya et al., 2014
Highlights Spiral fiber was utilized as biofilm carrier to get better adhesion of microorganisms Possible mechanism for nitrate removal was analyzed under Cr(VI) stress Excellent removal rates of nitrate and chromate were reached at low C/N ratio (0.9) The biofilm reactor achieved high removal rate (nitrate ≥ 99%, Cr(VI) ≥ 95%） The pH had impact on three procedure: denitrification, Cr(VI) reduction and DNRA
33
S0960-8524(17)30148-7 http://dx.doi.org/10.1016/j.biortech.2017.01.076 BITE 17604
To appear in:
Bioresource Technology
Received Date: Revised Date: Accepted Date:
2 November 2016 19 January 2017 22 January 2017
Please cite this article as: Zhai, S., Zhao, Y., Ji, M., Qi, W., Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor, Bioresource Technology (2017), doi: http://dx.doi.org/10.1016/ j.biortech.2017.01.076
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Simultaneous removal of nitrate and chromate in groundwater by a spiral fiber based biofilm reactor Siyuan ZHAI a, Yinxin ZHAO a,b*, Min JI a,b, Wenfang QI a a
School of Environmental Science and Engineering, Tianjin University, Tianjin
300350, China b
Tianjin Engineering Center of Urban River Eco-Purification Technology, Tianjin
300350, China Abstract: A spiral fiber based biofilm reactor was developed to remove nitrate and chromate simultaneously. The denitrification and Cr(VI) removal efficiency was evaluated with synthetic groundwater (NO3--N=50 mg/L) under different Cr(VI) concentrations (0~1.0 mg/L), carbon nitrogen ratios (C/N) (0.8~1.2), hydraulic retention times (HRT) (2~16 h) and initial pHs (4~10). Nitrate and Cr(VI) were completely removed without nitrite accumulation when the Cr(VI) concentration was lower than 0.4 mg/L. As Cr(VI) up to 1.0 mg/L, the system was obviously inhibited, but it recovered rapidly within 6 days due to the strong adaption and domestication of microorganisms in the biofilm reactor. The results demonstrated that high removal efficiency of nitrate (≥ 99%) and Cr(VI) (≥ 95%) were achieved at lower C/N = 0.9, HRT = 8 h, initial pH = 7, and Cr(VI) = 1.0 mg/L. The technology proposed in present study can be alternative for simultaneous removal of co-contaminants in groundwater. Key Words: Denitrification, Cr(VI) removal, Biofilm reactor, Groundwater *
Corresponding author: Yingxin Zhao. E-mail: [email protected] (Y.Zhao); Tel/Fax: +86 22 2740 6057
1
Introduction Groundwater is one of the main water resources, which is widely used as drinking water in most countries of the world. However, in recent years, the co-existence of different oxidized contaminants such as nitrate, sulfate, pesticide and chromate in natural water or wastewater becomes a common and growing problem (Li et al., 2016). In the nature, no matter the riverine system (Olivares et al., 2010), soil system (Katarzyna et al., 2016; Liu et al., 2015) and groundwater system (Babiker et al., 2004), the long-term combined contamination were more harmful to the environment and the toxicity of compound pollution could change the microbial community structure. Nitrate is the most ubiquitous chemical contaminant in groundwater due to excessive utilization of nitrogenous fertilizer in agriculture and discharge of domestic and industrial waste (Almasri et al., 2004; Wang et al., 2009; Chen et al., 2013).The presence of nitrate in groundwater is increasing worldwide, because it is not easily removed by conventional drinking water treatment (Glass et al., 1998). Nitrate can cause acute health problems when it is reduced into nitrite, which in turn can react with secondary or tertiary amines to produce carcinogenic nitrosamines. The US Environmental Protection Agency's standard has set the maximum contaminant level of nitrate at 10 mg/L and the same values are also proposed by China (Standards for drinking water quality, GB5749-2006).The conventional physicochemical methods used to eliminate nitrate from water are ion exchange (Bae et al., 2002), reverse osmosis (Schoeman et al., 2003) and electro-dialysis (Peel et al., 2003). In regards to 2
biological methods, biological denitrification is widely applied for nitrate removal from groundwater attributing to its high efficiency and low cost. Chromium, another environmental pollutant of interest in groundwater reservoirs, is one of the most abundant and toxic metals that cause the pollution of groundwater and soil due to its frequent industrial application, such as wood preservation, electroplating, alloy production and leather tanning (Sahinkaya et al., 2013). In the aquatic environment, chromium primarily exists in trivalent Cr(III) and hexavalent forms Cr(VI). Cr (VI) is highly soluble and mobile, which shows acute toxicity, mutagenicity and carcinogenicity in ecosystem (Somasundaram et al., 2009). While the relation between Cr(III) and Cr(VI) depends strongly on pH and the oxidative properties of the location. As a result, Cr(VI) reduction and subsequently immobilization of the generated Cr(III) is an effective method to remove and detoxify Cr(VI) (Li et al., 2016). The treatment of Cr(VI) polluted wastewater including chemical precipitation, electrochemical treatment, reverse osmosis and ion exchange (Barrera et al., 2012). However, the aforementioned methods present disadvantages such as high cost, energy consumption or production of chemical sludge, which makes them not suitable for removing low concentration of Cr(VI) in groundwater (Ganguli et al., 2002). Hence, biotransformation of Cr(VI) to the less toxic Cr(III) may be an alternative promising approach for groundwater treatment. Biological reduction of different oxidized contaminants to harmless or immobile forms is considered as a promising approach to solve compound pollution problems. Recently, Cr(VI) reduction by bacteria was found to have indivisible relationships 3
with the physiological electron acceptor (oxygen or nitrate) consumption (Somasundaram et al., 2009; He et al., 2015). In addition, some of previous studies have reported the simultaneous reduction of nitrate and chromate (Li et al. 2016; Chung et al., 2006; Miao et al., 2015; Sahinkaya et al., 2014). Chung et al. (2006) proposed that the simultaneous removal of nitrate and chromate in drinking water could be achieved using H2 as an electron donor in a H2-based membrane biofilm reactor. Sahinkaya et al. (2014) accomplished the removal of nitrate and Cr(VI)-containing wastewater in sulfur-based autotrophic denitrifying column bioreactors. Complete autotrophic denitrification was obtained at a nitrate loading rate of 0.1 g NO3--N/(L·d) and the influent Cr(VI) concentration below 0.5 mg/L. Lower removal rates of nitrate and chromate were attained, while the problem of secondary pollution by sulfur-based autotrophic denitrifying couldn’t be ignored. Recently, a solid substrate as an alternative to the soluble carbon source was developed, which could not only provide constantly reducing power for reduction of oxidized pollutants when attacked by microbial enzyme but also serve as support for biofilm formation and development. From the finding of Li, an up-flow packed-bed bioreactor was constructed to investigate the simultaneous removal of chromate. In this system, a biodegradable meal box was taken as carbon source and biofilm carriers, and complete denitrification and Cr(VI) reduction could be achieved with high Cr(VI) concentrations (2.0~50.0 mg/L) and longer HRT (>17h) (Li et al. 2016). However, continuous and stable release of organic carbon was also a problem by taking biodegradable meal box as solid carbon source. Therefore, more efforts should be 4
dedicated to develop a simple and efficient biofilm reactor to overcome the main disadvantages of existing technology, such as the high cost, low removal efficiency, secondary pollution and the complexity of reactor operation.. In this study, a spiral fiber carrier based biofilm reactor was developed to remove nitrate and chromate simultaneously. The simultaneous removal efficiencies of nitrate and chromate under different Cr(VI) concentrations, carbon and nitrate ratios (C/Ns), hydraulic retention times (HRTs) and initial solution pHs were investigated. In addition, the main Cr speciation and distribution around the biofilm were also analyzed. 2. Materials and methods 2.1 Experimental apparatus Fig.1 shows a set of lab-scale biofilm reactor used in this study. The main part of the reactor was an organic glass cylinders (diameter 300 mm, height 240 mm) with an effective volume of 7.5 L. The main reactor compartments consisted of a water collector (diameter 50 mm, height 200 mm), a water carrier and a water bath outer for heat preservation. The water collector was installed in the center of the cylinder, 108 holes (diameter 5 mm, distance between hole centers 20 mm) were drilled symmetrically in the collector wall. Each hole was filled with the carriers and the carriers bound on the collector. Peristaltic pumps were used to control the flow rate of influent and effluent. Synthetic groundwater was pumped from the influent tank into the biofilm reactor through the bottom inlet of the reactor. Influent flowed through the biofilm and effluent was drawn from the water collector. While the biofilm also 5
worked as a filter preventing suspended solids (SS) and other impurities into the treated water, therefore it could save a sedimentation tank or membrane filtration. 2.2 Chemicals and materials Synthetic contaminated groundwater (per liter of tap water) contained NaNO3, phosphate buffer solution and potassium dichromate. CH3OH was added as required carbon source to maintain the desired C/N. The original pH of the influent solution was normally about 7.0±0.2. All chemical reagents used were analytical grade without further purification. All solid reagents were weighed using analytical balance (Sartorius BS 124S, Germany) and liquid reagents were measured via pipettes with specific ranges. A mixed culture was originally collected from anaerobic sludge, which was obtained from the secondary sedimentation tank of Zhang-Gui-Zhuang Sewage Treatment Plant (Tianjin, China). In recent study, the anaerobic sludge collected from the same wastewater treatment plant contains some denitrifying bacteria which was proved by the results of the high-throughput sequencing. The carrier provided by Shijiazhuang green environmental protection technology co., Ltd. is composed of cotton, vinylon, polyvinylidene chloride and polypropylene fiber, and its shape is imitation of spiral aquatic plants type. The characteristics of the spiral fiber based biofilm reactor are strong adhesion of microorganisms, low cost and easy management. 2.3 Experiment procedure The biofilm reactor was set up to achieve the simultaneous removal of nitrate and 6
chromate under different operating conditions. The biofilm reactor was inoculated with anaerobic sludge and operated at 25-28 oC during the experiments. Biofilm formed gradually and a dark gray color was observed on the carrier within 10 days. During the whole operating periods, NO3--N concentration of influent was 50 mg/L. Then the biofilm reactor began to operate as indicated in Table 1. The biofilm reactor was operated for around 126 days under 4 different effect factors. In the first period, a low Cr(VI) concentration was kept at 0.1 mg/L for 12 days to enrich bacterial growth and promote biofilm formation. Then the influence of Cr(VI) concentrations (0.1, 0.2, 0.3, 0.4, 0.5 and 1.0 mg/L) on simultaneous nitrate and Cr(VI) removal was investigated. In phases 2, 3 and 4, the influent Cr(VI) concentration was determined at 1.0 mg/L. To study the effects of C/N and HRT on denitrification and Cr(VI) reduction, C/N was gradually decreased from 1.2 to 0.8, and the HRT was controlled at 16, 12, 8, 4, and 2 h. In the last period, the initial pHs of the system were conducted at 4, 7 and 10. During these procedures, the operating conditions of simultaneous nitrate and Cr(VI) removal were optimized. 2.4 Analysis method The influent and effluent water samples’ indicators of each cycle reaction were detected, including pH, CODCr, Cr(VI), Cr(III), NO3--N, NH4+-N and NO2--N. The pH in the influent and effluent was determined by a pH meter (Inesa PHSJ-3F, China). The chemical oxygen demand (COD) was measured using the dichromate method with HACH DR2800 (Rice et al., 2013). Cr(VI), NO3--N, NH4+-N and NO2--N were determined in accordance with standard methods. 7
A thermometer was set into the reactor to monitor the solution temperature during the experiments. 3. Results and discussion 3.1 Denitrification performance under different concentrations of Cr(VI) stress High Cr(VI) concentration in wastewater would conducted toxicity on activated sludge process, and the system was obviously affected as it was above 5 mg/L (Vaiopoulou et al., 2012). Comparing to wastewater, relatively lower Cr(VI) concentrations exist in groundwater environment, normally lower than 1.0 mg/L. Previous studies reported that 0.5 mg/L of Cr (VI) could impact on denitrification performance (Li et al. 2016). In the present study, different Cr(VI) concentrations (0.1, 0.2, 0.3, 0.4, 0.5 and 1.0 mg/L) were fed into the biofilm reactor during the first stage (1-38 d) to investigate the impact resistance of the system. The concentrations of NO3--N, NO2--N, NH4+-N and Cr(VI) in effluent under different influent Cr(VI) conditions were shown in Fig. 2. Denitrification performance under Cr(VI) stress was presented in Fig.2 (a). Almost complete denitrification was achieved when the concentrations of influent Cr(VI) were increased from 0.1 to 0.4 mg/L, which indicated that low concentrations of Cr(VI) had no obviously negative effects on denitrification. Increasing feed Cr(VI) concentration to 0.5 mg/L, the effluent nitrate concentrations increased to 1.39 mg/L, significantly responding the inhibiting effects of higher Cr(VI) on denitrification of the system. However, the nitrate removal efficiency recovered in two days and the nitrite concentration still remained at very low levels (≤ 0.075 mg/L). Further 8
increasing influent Cr(VI) concentration from 0.5 mg/L to 1.0 mg/L, obvious inhibiting effects were observed on denitrification. The nitrate removal rate decreased from almost 100% to 89.44%, and 0.78 mg/L NO2--N accumulated at the same time. The significant lag and inhibition of nitrate reduction herein were ascribed to the electron competition between nitrate and Cr(VI) (He et al., 2015). Another was that the Cr(VI) reduction process gave rise to reactive free radicals, such as Cr(V) and O2·, which further caused oxidative damage on DNA, RNA and proteins of the bacterial cells and severely influenced the enzyme synthesis and bacterial metabolism (Valko et al., 2005). After a period (about 7 days) operation, the microorganisms of the system may gradually produce a certain degree tolerance on higher Cr(VI) load through adaptation and domestication. Finally, the restoration of nitrate removal efficiency gradually improved and the effluent NO3--N concentration was below 0.5 mg/L. Bacteria reduce nitrate into nitrite with nitrate reductase enzymes (Nap or Nar), then nitrite can be transformed into ammonia through dissimilatory nitrate reduction to ammonia (DNRA) by nitrite reductase enzymes (Stolz et al., 2002). Therefore, the NH4 +-N variation (Fig. 2 (a)) was also analyzed during the whole operating periods of different Cr(VI) concentrations. At relatively low initial Cr(VI) level (0.1 mg/L and 0.2 mg/L), NH4+-N of the system accumulated to 2.53 mg/L. Then, the effluent ammonia concentrations were kept below 1.0 mg/L with increasing Cr(VI) concentration. The cytochromes of multiheme nitrite reductase (Nrf) can be readily oxidized by Cr(VI) thus rendering them inoperable (Stolz et al., 2011). As a result, the activity of the enzymes involved in DNRA will be suppressed by Cr(VI) adding. 9
The variations of influent and effluent Cr(VI) were plotted in Fig. 2(b). The effluent Cr(VI) rose obviously with increasing Cr (VI) load, but the system recovered after several cycles. During the reactor operation, the effluent Cr(VI) concentrations (≤ 0.05 mg/L) were even lower than the drinking water standard when the influent Cr(VI) concentration steadily increased up to 0.5 mg/L. As the influent Cr(VI) concentration was increased further, both denitrification and Cr(VI) removal capacity of the reactor decreased and effluent Cr(VI) reached around 0.18 mg/L at Cr(VI) concentration of 1.0 mg/L. Although a fluctuation of Cr(VI) decline occurred at each changed Cr(VI) load, Cr(VI) concentration finally reached below 0.05 mg/L based on the good adaption ability of the reactor. Cr(VI) is more stable under oxidizing conditions and Cr(III) is more stable under reducing conditions. The implementation of microbial chromium bioremediation has encountered, and anaerobic reduction of Cr(VI) can be mediated by several different microorganism. Peter (2012) reported three ways of biotransformation of nitrate and chromate contaminated environments: (1) competitive alternative electron acceptor; (2) co-metabolism; (3) induction of specific proteins and pathways involved in oxidation/reduction reactions. For the first way, the Cr(VI) reductase (ChrR) has more competition capability for electrons than nitrate reductase (Salamanca et al., 2013; Kourtev et al., 2009). In Fig.3 (a), effluent average CODCr concentration showed a significantly rise from 21.42 mg/L to 39.59 mg/L when the influent Cr(VI) was from 0.1 mg/L up to 0.2 mg/L. The decline of methanol utilization may be due to the inhibition of denitrification activity by Cr(VI) stress, and subsequently the amounts of dissolved 10
organic carbon (DOC) released exceeded what denitrification needed (Bae et al., 2002). However，it was worth noting that the effluent COD concentrations decreased with further increasing influent Cr(VI) concentration from 0.2 mg/L to 1.0 mg/L after a period operation. It proved that the biofilm reactor could tolerate higher chromium concentrations under heterotrophic conditions. The influent and effluent pH variation of the reactor were presented in Fig. 3 (b). During the overall operating periods under different Cr(VI) concentrations, no obvious effects of increasing influent Cr(VI) loading on effluent pH were observed. The initial pH maintained 7±0.2, the effluent pH was slightly above 7.5 due to the alkalinity generation of denitrification process (Equation 1). 6NO3-+5CH3OH→3N2+7H2O+5CO2+6OH-
(1)
3.2 Effect of C/N ratio on simultaneous nitrate and Cr(VI) removal
In this reactor, methanol was supplemented as carbon source. Through changing the methanol dosage, different C/N (1.2, 1.0, 0.9 and 0.8 mg/L) were studied in the biofilm reactor during the second stage (39~63 d) to investigate the impact resistance of the system. The concentrations of NO3--N, NO2--N and Cr(VI) in effluent under different C/N conditions were shown in Fig. 4. As seen in Fig.4(a), the effluent nitrate concentrations were quite low and almost complete denitrification was attained at relative higher C/N operation parameter (1.2~0.9). When C/N was above 0.9, the nitrite concentration in effluent was always below detection limit (0.005 mg/L NO2—N). Between days 54 and 59 (C/N = 0.8), nitrate and nitrite concentrations increased to 5.58 mg/L and 4.84 mg/L in the effluent, 11
respectively. This decline of denitrification efficiency may be due to limiting amount of methanol in the influent. At the same time, the Cr(VI) reduction process also responses to the shortage of the carbon source, and removal rate decreased to 80%. Sahinkaya did similar research about simultaneous bioreduction of nitrate and chromate under mixotrophic denitrification conditions (Sahinkaya et al., 2013). In their study, the optimal C/N ratio was 1.33 with combined heterotrophic and sulfur based autotrophic processes, and almost complete denitrification was achieved at influent NO3−–N and Cr(VI) concentrations of 75 mg/L and 10 mg/L, respectively. In comparison with Sahinkaya’s study, the biofilm reactor used in the present study performed better with lower C/N ratio of 0.9, indicating a cost advantage for its application. In order to recover denitrification and Cr(VI) reduction capabilities of the system, C/N was increased to 0.9 on day 59 of operation to create the optimum conditions for the microorganisms, thereby ensuring a stable and efficient operation of the biofilm reactor. Under the optimum operating condition, the denitrification and Cr(VI) reduction performance of the system was recovered in 3 days. 3.3 Effect of HRT on simultaneous nitrate and Cr(VI) removal
The impact of the key operating parameter-HRT on the nitrate and Cr(VI) removal performance of biofilm reactor was also investigated. With the presence of 50 mg/L nitrate, 210 mg/L COD and 1.0 mg/L Cr(VI) in the influent, the removal efficiencies of nitrate and COD at varying HRTs were shown in Fig.5. The average concentration of Cr(VI) and Cr(III) in effluent can be seen in Tab. 2. 12
As HRT decreased from 16 to 8 h, the removal efficiencies of nitrate, COD and Cr(VI) were in the ranges of 97~100%, 82.2~95.6% and 88~100%, respectively. There were no significant differences of pollutants removal efficiency as the HRT= 16, 12, 8 h, so the hydraulic retention time of 8 h could be identified as a suitable reaction parameters. Meanwhile, no accumulation of nitrite was detected, which indicated that the system was stable. The biological Cr(VI) reduction approach was controversial process for different researchers of different systems. The reduction pathway of Cr(VI) maybe reduced the biological toxicity to a certain extent, but not accomplish the thorough removal of Cr(VI) (Singh et al., 2011). In some studies, generated Cr(III) by Cr(VI) reduction were achieved almost 100% immobilization rate by biofilm reactor (Pan et al. 2014). While, Novotnik (2014) reported that Cr(VI) reduction mainly occurred in the intercellular portion. In our system, the Fig. 5(c) was shown that :one part of hexavalent chromium were removed completely; the other part of Cr(VI) transformed to Cr(III) and the Cr(III) concentrations were accumulated at the range of 0.02~0.21 mg/L when HRTs were higher than 8 h. Further decrease of HRT to 4 and 2 h, concomitantly, the contaminants removal rates were significantly decreased. Particularly from day 100 to 110, further decreasing HRT to 2 h led to a remarkable increase of nitrate (26.61 mg/L) and nitrite (9.18 mg/L) in the effluent and the denitrification rates decreased sharply to 60% (Fig.5(a) and Tab.2). While the COD removal rate collapsed rapidly from 90% to 70%. Besides, decreasing HRT led to an obvious decline of Cr(VI) removal rate to 48% 13
(Tab.2) and effluent total Cr = 0.82 mg/L (supported in the Fig.5c) was almost equal to influent Cr(VI) amount with little Cr(VI) transformation to Cr(III). To remove NO3--N and Cr(VI) effectively, the HRT was controlled at 8 h in the following studies. 3.4 Effect of initial pH on simultaneous nitrate and Cr(VI) removal
In order to recover denitrification and Cr(VI) reduction capability of the system and keep stabilization of the biofilm reactor for the next impact factor investigating, the HRT increased to 8 h and operating for 5 days. The pH of medium also played an important role in nitrate and Cr(VI) reduction. As shown in Fig.6, the system could effectively remove both nitrate and Cr(VI) at pH 7 and 10, which more than 95% of Cr(VI) removal efficiency was achieved and 99% nitrate removal efficiency was reached. The Cr(III) precipitation produced by Cr(VI) transformation was at a very low level (≤ 0.015 mg/L) during the operating periods under different initial pHs. Similarly, He et.al indicated that both Cr(VI) and nitrate could be removed effectively at pH from 8 to 9, in which more than 80% of Cr(VI) removal was achieved and nitrate was depleted (He et al., 2015). In addition, effluent pHs were steadily in the range of 7.5 to 8.0. As the initial pH was conducted to 4, the acidic condition distinctly defeated the nitrate and Cr(VI) reduction ability. While the NO3--N concentration reached 1.76 mg/L and Cr(VI) accumulated to 0.15 mg/L. The growth and activities of microorganism would be suppressed significantly at the condition of strong alkaline (pH > 10) or acidic (pH < 6) which distinctly defeated the Cr(VI) and nitrate reduction ability (He et al., 2015). 14
Generally, the pH may be a factor affecting another pathway DNRA. Some previous investigations reported that the optimum pH was 6.5 and 7.5 for NO2--N and NO3--N reduction, and the higher DNRA was generally observed in alkaline environments (Stevens et al. 1998; Rütting et al. 2011; Zhang et al. 2015).With the system changing from acidic to alkaline condition in our system, the obviously increasing NH4+-H concentration was found by the DNRA pathway. This phenomenon was consistent with the findings of Zhang (2015), who stated that the DNRA process was negligible under acidic condition(pH 4.7), but was mor prevalent in neutral (pH 6.2) and alkaline (pH 8.2) environments. 3.5 Comparison of nitrate and Cr(VI) removal with different systems
Tab.3 presented a comparison between the reactor used in this study and other system used for the simultaneous removal of nitrate and Cr(VI), including the information about the main operational parameters and removal efficiencies of each system. It can be seen from Tab.3 that the biofilm reactor in the present study has significant advantages. The obvious superiority lies in high removal efficiency of contaminants (≥ 99% nitrate and ≥ 95% Cr(VI)), lower carbon and nitrogen ratio (0.9) and shorter hydraulic retention time (8 h). The operational costs of this system are mainly related to carbon source, therefore the use of a low carbon to nitrogen ratio reduces the operational costs for its application in the treatment of co-contaminants present in groundwater. It is apparent that the biofilm reactor is favorable for treatment of nitrate and Cr(VI) co-contaminated groundwater. 15
4. Conclusions The simultaneous removal of nitrate and Cr(VI) was achieved by a spiral fiber based biofilm reactor. Low Cr(VI) concentration (≤0.4 mg/L) had no obviously negative effects on nitrate removal. The denitrification performance was inhibited as the Cr(VI) concentration increased to 1.0 mg/L, but the restorations of nitrate and Cr(VI) removal efficiency gradually improved more than 99% and 95% after a period operation. The optimum C/N, HRT and initial pH were observed at 0.9, 8 h and pH=7. It indicated that the method had promising application for treatment of nitrate and Cr(VI) co-contaminated groundwater.
16
Acknowledgments： ： Authors are grateful to the financial support from the National Key Technology Research and Development Program (Project No. 2015BAL04B03, China). and Critical Patented Projects in the Control and Management of the National Polluted Water Bodies (Project No. 2015ZX07203-011, China)
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Figure Captions Fig.1. Experimental apparatus Fig.2. Denitrification and Cr(VI) removal under different concentrations of Cr(VI) Fig.3. Variations in effluent CODCr concentrations and pH under different concentrations of Cr(VI) Fig.4. Denitrification and Cr(VI) removal under different C/N Fig.5. The biofilm reactor removal efficiency under different HRTs Fig.6.The impact of initial pH on system denitrification and Cr(VI) reduction performance
22
6 11
b 7 9
a
3
5
10
4 2 1
12
8
Fig.1 Experimental apparatus (1.reactor; 2. water bath outer; 3.water collector; 4.carriers; 5.wastewater inlet; 6.treated water outlet; 7.overflow pipe; 8. constant temperature circulating water inlet; 9. constant temperature circulating water outlet; 10,11. peristaltic pump; 12. influent tank; a. influent pipe; b. effluent pipe.)
23
6
-
NO3 -N
5 N concentration / (mg/L)
1.0
Influent Cr(VI) -
+
NO2 -N
NH4 -N
4
0.5
3 2
0.0
1 0
0
10
20
30
Influent Cr(VI) /(mg/L)
(a)
-0.5 40
Time/(d)
0.30
Influent Cr(VI)
Influent Cr(VI)/(mg/L)
Effluent Cr(VI)
0.25
0.9 0.20 0.15
0.6
0.10 0.3 0.05 0.0
0
10
20
30
Effluent Cr(VI)/(mg/L)
(b)1.2
0.00 40
Time/(d)
Fig.2 Denitrification and Cr(VI) removal under different concentrations of Cr(VI)
24
(a)
80
Influent COD Cr=205~215 mg/L
Effluent CODCr / (mg/L)
Effluent COD Cr
60
40
20
0
0.1
0.2
0.3
0.4
0.5
1.0 6
Influent Cr(VI)/(mg/L)
1.0
Influent Cr(VI) / (mg/L)
9.0
Influent Cr(VI) pH variation
8.5
0.8
8.0
0.6
7.5
Initial pH=7±0.2
0.4
7.0
0.2 0.0
pH variation
(b) 1.2
6.5
0
10
20
30
6.0 40
Time/(d)
Fig.3 Variations in effluent CODCr concentrations and pH under different concentrations of Cr(VI)
25
(a)
8
1.4
C/N ratio NO2--N
6
1.2 1.2
4
1.0 0.9
1.0
0.8
2
0
0.9
40
44
48
52
56
0.8
60
64
C/N variation
N concentration /(mg/L)
NO3--N
0.6
Time/(d)
(b)0.25
1.4
C/N ratio Effluent Cr(VI)
1.2 1.2
0.15 0.10
1.0
0.9
1.0
0.9 0.8
0.8
0.05 0.00
40
44
48
52
56
60
64
C/N variation
Effluent Cr(VI) /(mg/L)
0.20
0.6
Time/(d)
Fig.4 Denitrification and Cr(VI) removal under different C/Ns
26
N concentration / (mg/L)
25
HRT NO3 -N
20 + NH4 -N
NO2 -N
16
20
12
15 8
10
4
5 0
70
80
90
100
110
HRT variation /(h)
(a) 30
0
Time/(d)
75
20
HRT Effluent CODCr
16
60
12
45
8
30
30mg/L 15 0 60
70
80
90
100
110
4
HRT variation / (h)
CODCr concentration / (mg/L)
(b) 90
0
Time / (d)
Cr(VI) Cr concentration / (mg/L)
20
HRT Cr(III)
0.8
15
0.6 10 0.4 5
0.2 0.0
70
80
90
100
110
HRT variation / (h)
(c) 1.0
0
Time / (d)
Fig.5 The biofilm reactor removal efficiency under different HRTs
27
contaminant concentrations / (mg/L)
4
NO 3 - -N
NO 2 - -N
NH 4 + -N
Cr(VI)
3
2
1
0
41
2
7
initial pH
3
10
Fig.6 The impact of initial pH on system denitrification and Cr(VI) removal performance
28
Table Captions Tab.1. Operating periods of biofilm reactor Tab.2. Nitrate and Cr(VI) reduction activity of different HRTs Tab.3. Simultaneous removal of nitrate and chromate in different reaction system
29
Tab.1 Operating periods of biofilm reactor Periods Operating dates/(d) NO3--N/(mg/L) Cr(VI)/(mg/L) C/N HRT/(h) Initial pH
1
2
3
4
1-38 50 0.1,0.2,0.3,0.4,0.5,1.0 1.2 8 7
39-63 50 1 0.8,0.9.1.0,1.2 8 7
64-110 50 1 0.9 2,4,8,12,16 7
115-136 50 1 0.9 8 4,7,10
30
Tab.2 Nitrate and Cr(VI) reduction activity of different HRTs HRT(h) Nitrate removal rates (%)
16
12
8
4
2
100
100
99
94
60
Effluent Cr(VI) (mg/L)
0.036±0.02 0.041±0.01 0.048±0.05
0.32±0.12 0.52±0.24
Effluent Cr(III) (mg/L)
0.009±0.01 0.012±0.04 0.008±0.07
0.18±0.10 0.24±0.08
31
Tab.3 Simultaneous removal of nitrate and chromate in different reaction system
Reaction system Biofilm reactor Expanded granular sludge bed reactor Up-flow packed-bed bioreactor Autotrophic column bioreactor Heterotrophic column bioreator
NO3--N (mg/L)
Cr(VI) (mg/L)
50
1.0
200
10
100
2.0
75
2.0
75
2.0
Operating conditions (Methnol)C/N=0.9, HRT=8 h Liquid up-flow velocity =2.0 m/h (BMB)COD=400 mg/L HRT=17 h, (Methnol)C/N=2.85, HRT=12h (Methnol)C/N=3.41, HRT=12h
32
Nitrate/Cr(VI) removal rate(%) ≥99/95 ≥99.5/95 ≥99/95 ≥64/≥99/-
Reference This study Miao et al., 2015 Li et al., 2016 Sahinkaya et al., 2014 Sahinkaya et al., 2014
Highlights Spiral fiber was utilized as biofilm carrier to get better adhesion of microorganisms Possible mechanism for nitrate removal was analyzed under Cr(VI) stress Excellent removal rates of nitrate and chromate were reached at low C/N ratio (0.9) The biofilm reactor achieved high removal rate (nitrate ≥ 99%, Cr(VI) ≥ 95%） The pH had impact on three procedure: denitrification, Cr(VI) reduction and DNRA
33