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Simultaneous removal of perchlorate and nitrate in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy
Chemical Engineering Journal 284 (2016) 1008–1016
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Simultaneous removal of perchlorate and nitrate in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy Mengchun Gao a, Sen Wang b, Yun Ren b, Chunji Jin a, Zonglian She a,⇑, Yangguo Zhao a, Shiying Yang a, Liang Guo a, Jian Zhang b, Zhiwei Li b a b
Key Lab of Marine Environment and Ecology, Ministry of Education, Ocean University of China, Qingdao 266100, China College of Environmental Science and Engineering, Ocean University of China, Qingdao 266100, China
h i g h l i g h t s
0
S - and H2-autotrophy for NO3 and ClO4 reduction were investigated in a reactor. Autotrophic denitrifying process favored over autotrophic perchlorate reduction. +
0
H from S autotrophy process was consumed by electrochemical H2 autotrophy.
Some autotrophic denitrifier can use both NO3 and ClO4 as electron acceptor. Microbial communities at different positions in the reactor were analyzed.
a r t i c l e
i n f o
Article history: Received 23 June 2015 Received in revised form 24 September 2015 Accepted 25 September 2015
Keywords: Perchlorate Nitrate Sulfur autotrophy Electrochemical hydrogen autotrophy S0 particle
a b s t r a c t The removal of perchlorate (ClO 4 ) and nitrate (NO3 ) from drinking water was investigated in a combined reactor of sulfur autotrophy (S-compartment) and electrochemical hydrogen autotrophy (H-compartment). The removal efficiencies of NO and ClO in the S-compartment and 3 4 H-compartment were affected by hydraulic retention time (HRT) and current intensity, respectively. The sulfur- and hydrogen-autotrophic denitrifying process favored over the process of sulfur- and hydrogen-autotrophic perchlorate reduction in the combined reactor. The longer HRT could lead to sulfur (S0) disproportionation due to the increase of the contact time between water and S0 particle. The H+ generated from S-compartment could be reduced as H2 by electrochemical process in H-compartment, and the generated H2 as an electron donor was utilized to reduce NO 3 and ClO4 by hydrogen autotrophic reduction. The oxidation reduction potential (ORP) in the effluent from S-compartment and H-compartment were below 180 mV, suggesting good anaerobic conditions for the reduction of NO 3 and ClO 4 in the combined reactor. Some sulfur- and hydrogen-autotrophic denitrifying bacteria could use both NO3 and ClO4 as electron acceptors in the combined reactor. The DGGR profile illustrated that some variations were found in the microbial community at different locations of the combined reactor. Ó 2015 Elsevier B.V. All rights reserved.
1. Introduction Perchlorate (ClO 4 ) has been widely used in solid rocket fuels, missiles, explosives, pyrotechnics, and fireworks [1–3]. ClO 4 contamination in surface water and groundwater has become a significant environmental concern in recent years. ClO 4 can disrupt thyroid function by interfering with body’s intake of iodine, which inhibits the production of thyroid hormone [4]. Nitrate (NO 3 ) is ⇑ Corresponding author at: Key Lab of Marine Environment and Ecology, Ministry of Education, Ocean University of China, Qingdao 266100, China. Tel.: +86 532 66786351; fax: +86 532 66782810. E-mail address: [email protected] (Z. She). http://dx.doi.org/10.1016/j.cej.2015.09.082 1385-8947/Ó 2015 Elsevier B.V. All rights reserved.
often a co-contaminant with ClO 4 in surface water and groundwater, which mainly comes from agricultural runoff and wastewater discharge. The consumption of water contaminated by NO 3 can cause methemoglobinemia in infants, malformation and mutation [5]. The current technologies for ClO 4 and NO3 removal from drinking water and groundwater include ion exchange, photocatalytic reduction, membrane technology, absorption and biological reduction [6–13]. Ion exchange and biological reduction are regarded as the most cost–effective technologies for the treatment of ClO 4 and NO 3 contaminated water. The major disadvantage of ion exchange is that it can generate ClO 4 - and NO3 -laden brine, which requires specialized disposal and/or treatment. Biological reduction may
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M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016 completely transform ClO and N2, 4 and NO3 into innocuous Cl respectively. Some biological processes using different organic substrates as electron donors have been proved to be highly effec tive for ClO 4 and NO3 removal [14–17]. However, the residual organic substrate in the effluent can stimulate microbial growth in water distribution system and contribute to the formation of potentially toxic trihalomethanes during disinfection by chlorination [18]. To settle the above-mentioned problems, autotrophic reduction of ClO 4 and NO3 has been carried out by using inorganic matters as electron donors [19–21]. Hydrogen (H2) autotrophic reduction of ClO 4 and NO3 has been evaluated in some continuous flow bioreactors [19,22]. Although H2 is a good energy substrate for ClO 4 and NO3 reduction, it is difficult to handle in bulk quantities and is publicly perceived to represent a significant disaster threat due to its inherent explosive nature. Granular sulfur (S0) in packed bed bioreactors has been successfully used as an electron donor in the autotrophic reduction of ClO 4 and NO3 [20,23–26]. As granular S0 is insoluble in water, it can provide a slow-release supply of electron on demand and offer low expense cost as well as low maintenance cost. The stoichiometric equations for the S0 auto trophic reduction of ClO 4 and NO3 are respectively shown as follows [24,26]:
DC electrical source
+
_
Anode Carbon particle
Cathode
H2
10 cm
H-Part
H1 10 cm
S5 S4
þ
! 5:7 H þ
2:9 SO2 4
þ Cl þ 0:46 C5 H7 O2 N
ð1Þ
10 cm Sampling port 10 cm
S-Part
S3 10 cm
S2
2:9 S0 þ ClO4 þ 3:3 H2 O þ 1:8 CO2 þ 0:46 HCO3 þ 0:46 NHþ4
Effluent
S1
9 cm 1 cm S0 particle
1:1 S0 þ NO3 þ 0:76 H2 O þ 0:4 CO2 þ 0:08 NHþ4 ! 1:28 Hþ þ 1:1 SO2 4 þ 0:5 N2 þ 0:08 C5 H7 O2 N
Influent tank
ð2Þ
The main disadvantage for sulfur autotrophic ClO 4 and NO3 + reduction is the formation of sulfate (SO2 4 ) and H in the effluent. Limestone is often used as a low-cost alkalinity source, whereas it can increase the hardness of the effluent due to the release of Ca2+ during limestone dissolution. To make up for the deficiency of sulfur autotrophic ClO 4 and NO3 reduction, a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy was used to remove ClO 4 and NO3 in the present study. In the combined reactor, the H+ generated from sulfur-autotrophy ClO 4 and NO 3 reduction could be consumed by the bioelectrochemical hydrogen-autotrophy ClO and NO reduction to achieve 4 3 neutralization, and the SO2 concentration in the effluent can be 4 controlled by adjusting the ClO 4 and NO3 loading of sulfur autotrophy and electrochemical hydrogen autotrophy. The main objectives of the present study were (1) to evaluate the feasibility of ClO 4 reduction by sulfur- and hydrogenautotrophic denitrifying bacteria; (2) to investigate the effect of hydraulic retention time (HRT) and current intensity on ClO 4 and NO 3 reduction in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy; (3) to analyze the variation of microbial community along the flow path in the combined reactor.
stainless column. The space between the cathode and the anode in the H-compartment was filled with carbon particle with 3.0– 4.0 mm in diameter. Before the synthetic wastewater was fed to the combined reactor, it was firstly deoxidized by injecting nitrogen gas in the influent tank. A peristaltic pump was used to feed the influent into the combined reactor, and direct current was supplied to the H-compartment by a silicon rectifier. The Chemical processes of NO 3 and ClO4 reduction in the S-compartment and H-compartment is shown in Fig. 2. The water sampling ports were mounted at the height of 1 cm (S1), 10 cm (S2), 20 cm (S3), 30 cm (S4), 40 cm (S5), 50 cm (H1) and 60 cm (H2) from the bottom of the combined reactor. In addition, the S0 granules in the S-compartment were not replenished during the whole operational periods.
2. Materials and methods
2.2. Inoculated sludge and synthetic wastewater
2.1. Experimental set-up
The inoculated sludge was taken from an anaerobic tank of Lichunhe wastewater treatment plant in Qingdao city, China. The synthetic wastewater fed to the combined reactor was prepared daily by adding NaNO3, NaClO4H2O, and nutrient medium to the tap water. The compositions of nutrient medium were shown as follows: NH4HCO3 0.410 g/L, NaHCO3 2.700 g/L, MgSO47H2O 0.100 g/L, Ca(OH)2 0.005 g/L, and trace element solution 1 mL/L. The trace element solution consisted of the following components (g/L): Na2SeO35H2O 0.1, H3BO3 0.05, MnSO4H2O 0.415, NiSO46H2O 0.113, FeSO47H2O 2.8, Na2WO42H2O 0.5, CoSO412H2O 2.36, KAl (SO4)212H2O 0.175, EDTA 1.0, ZnSO47H2O 0.106, (NH4)6Mo7O24H2O 0.05, and CuSO45H2O 0.157.
The schematic diagram of an experimental set-up is shown in Fig. 1, which consists of sulfur autotrophy (S-compartment) and electrochemical hydrogen autotrophy (H-compartment). The Scompartment was a cylindrical plexiglass column with 7.5 cm in diameter and 40 cm in height, which was filled with S0 granules of 3.0–4.0 mm in diameter. A cylindrical stainless column with 7.5 cm in diameter and 25 cm in height was used as the cathode in the H-compartment. A graphite rod with 2.5 cm in diameter and 25 cm in height was used as the anode in the Hcompartment, which was located in the center of the cylindrical
Influent pump
Fig. 1. Schematic diagram of an experimental set-up.
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2H2O +2e
ClO4NO3-
Electrolysis
2OH+ Electrochemical reduction H2 H2O 20 SO4 + 2e S -2e-6eClO4 Cl- + N2 Cl- + N2 + 2H+ NO3- H2 autotrophic bacteria S0 autotrophic bacteria S - c o m p a r t m e nt
H-compartment
Fig. 2. Chemical processes of NO 3 and ClO4 reduction in the S-compartment and H-compartment.
Table 1 Operational conditions of reactor #1. Operational stages Operational time (d)
I 0–20
II 20–32
III 32–50
IV 50–64
V 64–84
VI 84–104
VII 104–122
VIII 122–168
Influent NO 3 -N concentration (mg/L) Influent ClO 4 concentration (mg/L) HRT (h) Current intensity (mA) Voltage (V)
28 0 12 10 2
28 0 8 10 2
28 0 4 10 2
28 0 2 10 2
28 0 1 10 2
28 0 0.5 10 2
28 0 0.5 20 4
0 124.38 0.5 20 4
Table 2 Operational conditions of reactor 2#. Operational periods Operational days (d)
I 0–38
II 38–58
III 58–80
IV 80–106
V 106–126
VI 126–152
VII 152–170
VIII 170–188
IX 188–214
X 214–238
XI 238–252
Influent ClO 4 concentration (mg/L) Influent NO 3 -N concentration (mg/L) HRT (h) Current intensity (mA) Voltage (V)
2 20 12 10 2
10 3 8 10 2
10 30 5 10 2
10 30 3 10 2
10 30 2 10 2
10 30 1 10 2
10 30 0.5 10 2
10 30 0.5 15 3
10 30 0.5 25 5
10 30 0.5 35 7
10 30 0.5 45 9
2.3. Operational procedure
3. Results and discussion
Two identical experimental set-ups were prepared in the present study. The first reactor (#1) was used to evaluate the probability of ClO 4 reduction by sulfur- and hydrogen-autotrophic denitrifying bacteria. The second reactor (#2) was used to investi gate the effect of HRT and current intensity on ClO 4 and NO3 reduction in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. The operational procedures of reactor #1 and #2 were summarized in Tables 1 and 2, respectively.
3.1. Feasibility assessment of perchlorate reduction by autotrophic denitrifying bacteria in the combined reactor
2.4. Analytical methods The water samples were filtered using a 0.22 lm filter membrane and then stored in a refrigerator at 4 °C for further analysis. The oxidation reduction potential (ORP) was measured by a portable ORT meter. A portable pH meter (Model 6010, WTW, Germany) was used to monitor pH in the influent and effluent. NO 3 , ClO4 , and SO2 were determined by an ion chromatography (ICS-300, 4 Dionex, USA). In order to evaluate the distribution of microbial community along the flow path of reactor #2, the biofilm samples were obtained at the height of 10 cm, 20 cm, 30 cm, 40 cm, 50 cm and 58 cm from the bottom at the end of the experiment (day 252), respectively. The DNA extraction, denaturing gradient gel electrophoresis (DGGE) analysis and sequencing were performed according to the methodology reported by Wang et al. [27].
Most perchlorate-reducing bacteria can utilize NO 3 as an alternative electron acceptor under anoxic conditions [2,28]. Some denitrifying bacteria can reduce ClO 4 using nitrate reductase enzymes as well as specialized perchlorate reductases [29,30]. However, some researchers have also found that the common denitrifying bacteria cannot use chlorate (ClO 3 ) or ClO4 as an electron acceptor [28,31–33]. In order to valuate the feasibility of ClO 4 reduction by autotrophic denitrifying bacteria, the ClO 4 reduction by sulfur- and hydrogen-autotrophic denitrifying bacteria was investigated in the combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy at different conditions (Table 1). According to Eqs. (1) and (2), the complete reductions of NO 3 and ClO4 per mol need to transfer 5 mol e and 8 mol e, respectively. Considering the equilibrium of electron transfer, the influent concentrations of NO 3 -N in stages I–VII and ClO4 in stage VIII were 28 and 124.38 mg/L, respectively, corresponding to 2 mmol/L NO 3 and 1.25 nmol/L ClO 4 in the influent. Fig. 3 shows the variation of NO (or ClO concentration in the effluent from 3 -N 4) S-compartment and H-compartment in different operational stages. In the operational stages I–V, the NO 3 -N in the effluent from both S-compartment and H-compartment at steady state was nearly not detected under the conditions of HRT varying from 12 to 1 h and 10 mA current. When the HRT was shortened to 0.5 h in the operational stage VI, the average NO 3 -N concentrations in
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M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016
Operational stage
Operational stage I
III
IV
V
VI
VIII
VII
I
II
III
IV
V
VI
VII VIII
IX
X
XI
11
Influent NO 3--N
Effluent NO 3--N from S-com
Effluent NO 3--N from H-com
Inffluent ClO 4-
Effluent ClO 4- from S-com
Effluent ClO 4- from H-com
10 9
2.0
ClO4- concentration (mg/L)
ClO4- or NO3--N concentration (mmol/L)
2.5
II
1.5
1.0
Influent ClO 4-
8
Effluent ClO 4- from S-com
7
Effluent ClO 4- from H-com
6 5 4 3 2
0.5
1 0 0.0
0 0
20
40
60 80 100 Operational time (d)
120
140
20
40
60
80
160
100 120
140
160 180
200
220 240
IX
X
Operational time (d) I
Fig. 3. Nitration reduction (or perchlorate reduction) by autotrophic denitrifying bacteria in the combined reactor. S-compartment, S-com; H-Compartment, H-com.
II
III
IV
V
VI
VII VIII
XI
35
3.2. Simultaneous removal of ClO 4 and NO3 in different operational stages
Some changes were made for the combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. These changes included the increase in current intensity from 10 to 45 mA and the decrease in HRT from 12 to 0.5 h (Table 2). The NO 3 and ClO4 concentrations in the effluent were consistent for at least 10 days in an operational stage before the next operational stage began in the combined reactor. Fig. 4 shows the removal effi ciencies of ClO 4 and NO3 in the combined reactor in different operational stages. In the operational stage I, the reactor was fed 20 mg/L NO 3 -N and 2 mg/L ClO4 under the conditions of 12 h HRT and 10 mA current. No NO in the effluent from 3 S-compartment was detected at steady state, suggesting the H-compartment did not participate in the process of NO 3 reduction. The average ClO 4 concentration in the effluent from S-compartment at steady states was 0.49 ± 0.01 mg/L in the operational stage I, whereas no ClO 4 was detected in the effluent from H-compartment. In the operational stages II–VI, the reactor was fed 30 mg/L NO 3 -N and 10 mg/L ClO4 under the conditions of 10 mA current and HRT varying from 8 to 1 h. The effluent NO 3 from the S-compartment at steady state was not detected in operational stage II–VI, which illustrated that the influent NO 3 could be
-
NO3 -N concentration (mg/L)
30
the effluent from S-compartment and H-compartment at steady states were 8.86 ± 0.03 mg/L and 2.40 ± 0.01 mg/L at 10 mA current, respectively. In order to increase the denitrification efficiency of H-compartment, the current was increased to 20 mA for producing more H2 in the stage VII. As shown in Fig. 3, the average NO 3 -N concentrations in the effluent from S-compartment and H-compartment at steady states was 5.87 ± 0.02 mg/L and 0 mg/L in the operational stage VII, respectively. In the stage VIII, the concentrations of ClO 4 and NO3 in the influent were 124.38 and 0 mg/L, respectively. The average ClO 4 concentration in the effluent from S-compartment and H-compartment at steady states was 98.63 ± 0.22 mg/L and 94.53 ± 0.31 mg/L in the stage VIII, respectively. Compared with the NO 3 reduction in stage VII, the sulfur- and hydrogen-autotrophic denitrifying bacteria could partially reduce ClO 4 in the present study, suggesting some auto trophic denitrifying bacteria could use both NO 3 and ClO4 as electron acceptors or only use NO 3 as an electron acceptor.
Influent NO 3--N
25
Effluent NO 3--N from S-com
20
Effluent NO 3--N from H-com
15 10 5 0 0
20
40
60
80
100
120 140
160 180
200 220
240
Operational time (d) Fig. 4. Removal efficiencies of ClO 4 and NO3 -N in the combined reactor in different operational stages. S-compartment, S-com; H-Compartment, H-com.
removed in the S-compartment at 1–8 h HRT. The average effluent ClO 4 concentrations from S-compartment at steady states were 2.03 ± 0.06, 1.06 ± 0.02, 1.00 ± 0.01, 1.51 ± 0.03 and 2.04 ± 0.05 mg/L at the HRT of 8, 5, 3, 2 and 1 h, respectively. However, no ClO 4 in the effluent from H-compartment at steady states was detected in operational stage II–VI. In the operational stage VII, the average NO 3 -N and ClO4 concentrations in the effluent from S-compartment increased to 8.92 ± 0.06 and 4.98 ± 0.04 mg/L as the HRT decreased to 0.5 h, respectively. Similarly, the average NO and ClO concentrations in the effluent from 3 -N 4 H-compartment increased to 2.48 ± 0.03 and 3.99 ± 0.10 mg/L in the operational stage VII, respectively. Compared to the stage VI, the NO concentrations in the effluent from 3 -N and ClO4 H-compartment increased in the stage VII due to the shortage of HRT, which was peculated that H2 as the electron donor in H2-compartment could be inadequate for the reduction of NO-N and ClO 4 in stage VII. In order to enhance the hydrogen autotrophic reduction of NO 3 and ClO4 in the H-compartment, the current intensity increased from 10 to 45 mA with the conversion of the operational stage from V II to XI. Compared with the operational stage VII, the average effluent NO 3 and ClO4 concentrations from S-compartment had no obvious variation, suggesting the removal efficiencies of NO 3 and ClO4 were related to HRT in the S-compartment. However, No NO was detected from 3 H-compartment in the operational stages VII–XI, and the average effluent ClO 4 from H-compartment decreased from 3.99 ± 0.10 to
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M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016
1.49 ± 0.08 mg/L with the increase in current from 10 to 45 mA. As H2 as the electron donor in the H-compartment originated from the electrochemical reduction of H+ from the S-compartment or water electrolysis process, the H2 production was closely related to the current intensity in the H2-compartment. The present results suggested that the removal efficiencies of NO 3 and ClO4 in the H-compartment could be affected by the variation of current intensity. 3.3. Variation of ClO 4 and NO3 concentrations along the flow path of the combined reactor In order to investigate the relative progress of ClO 4 and NO3 reduction in the combined reactor (#2), the variations of ClO 4 and NO 3 concentrations at steady state were analyzed along the flow path of the combined reactor. The concentrations of ClO 4 and NO 3 decreased along the flow path of the combined reactor in different operational stages. This phenomenon could be explained that sulfur- (or hydrogen-) autotrophic reduction bacteria and denitrifying bacteria coexisted in S-compartment (or H-compartment) or that some autotrophic bacteria could simulta neously utilize NO 3 and ClO4 as electron acceptors. As shown in Fig. 5, no NO3 was detected in the S4 sampling port under the conditions of 10 mA current and HRT varying from 3 to 1 h, whereas the effluent ClO 4 concentration from the S4 sampling port increased from 4 to 6 mg/L with the decrease in HRT from 3 to 1 h. No ClO 4 in the effluent was found in the sampling port of H1, H2 and H2 at HRT of 3, 2 and 1 h, respectively. The NO 3 -N and ClO 4 concentrations from the sampling port H2 were 3 and
HRT =3h, I=10mA HRT =2h, I=10mA HRT =1h, I=10mA HRT =0.5h, I=10mA
ClO4- concentration (mg/L)
10
8
HRT =0.5h, I=15mA HRT =0.5h, I=25mA HRT =0.5h, I=35mA
6
4
2
5 mg/L under 10 mA current intensity and 0.5 h HRT, respectively. The ClO 4 concentration from sampling port H2 decreased with the increase in current from 10 to 35 mA, whereas no NO 3 from sampling port H1, H1 and H2 was detected under the conditions of 15–35 mA current intensity and 0.5 h HRT. The abovementioned results showed that the processes of autotrophic perchlorate reduction and denitrification could occur in parallel when sufficient substrates were present in the combined reactor. Com pared to NO 3 reduction, a lag in ClO4 reduction was found in the S-compartment and H-compartment of the combined reactor. According to the reports of Ontiveros-Valencia et al. [34,35], the present results could be explained that some microorganisms pre fer NO 3 over ClO4 as electron acceptor, which delayed the onset of sulfur- and hydrogen-autotrophic ClO 4 reduction. The sulfur- (or hydrogen-) autotrophic ClO 4 reduction might begin and occur simultaneously with the sulfur- (or hydrogen-) autotrophic denitrification before NO 3 was completely removed from the influent containing NO 3 and ClO4 . In other words, some sulfur- (or hydrogen-) autotrophic denitrifying bacteria in the present study might not utilize ClO 4 as an electron acceptor, and others could remove both ClO 4 and NO3 or retain the capacity for ClO4 reduc tion when exposed to both ClO4 and NO3 . Ju et al. [20] reported 0 that the addition of NO 3 to the S -oxidizing perchlorate reduction bacteria delay the onset of ClO 4 reduction until NO3 was partially consumed. 3.4. Variation of SO2 4 concentration in the effluent from the combined reactor 0 The coupling of ClO 4 and NO3 reduction with S oxidation can concomitantly produce SO2 in S-compartment. According to Eqs. 4 (1) and (2), the SO2 4 production from ClO4 and NO3 reduction in S-compartment is stoichiometrically 2.80 mg SO2 4 /mg ClO4 and 7.54 mg SO2 /mg NO -N, respectively. As shown in Fig. 6, the aver4 3 age effluent SO2 concentration at steady states was 4 265.24 ± 4.84 mg/L from S-compartment when the influent with 20 mg/L NO 3 -N and 2 mg/L ClO4 was fed to the combined reactor at 12 h HRT in the operational stage I. The SO2 4 production in the operational stage I was much higher than the theoretical SO2 4 pro duction (156.4 mg/L) from NO 3 and ClO4 reduction. The production of excessive SO2 4 in the operational stage I showed that the sulfur disproportionation could happen in the reactor as follows:
þ 4S0 þ 4H2 O ! 3H2 S þ SO2 4 þ 2H
ð3Þ
0 S1
S2
S3
S4
S5
H1
H2
Operational stage
Sampling port
I
III
IV
280
30
HRT =3h, I=10mA HRT =2h, I=10mA HRT =1h, I=10mA HRT =0.5h, I=10mA HRT =0.5h, I=15mA HRT =0.5h, I=25mA HRT =0.5h, I=35mA
25 20
V
VI
VII VIII
IX
X
XI
Influent Effluent from S-compartment Effluent from H-compartment
260 240 SO42- concentration (mg/L)
NO3 --N concentration (mg/L)
II
15 10
220 200 180 160 140 120
5
100
0 S1
80
S2
S3
S4
S5
H1
H2
Sampling port Fig. 5. Variation of ClO 4 and NO3 -N concentrations along the flow path in the combined reactor.
0
20
40
60
80
100 120 140 160 180 Operational time (d)
200 220
240
Fig. 6. Variation of SO2 4 concentration in the effluent from the combined reactor in different operational stages.
M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016 Some researchers have reported that ClO 4 and NO3 reduction 0 0 using S as the electron donor coupled with S disproportionation in batch experiment or continuous flow experiment [24,36–37]. In the operational stages II–XI, the reactor was fed 30 mg/L NO 3 -N and 10 mg/L ClO 4 in the influent, and the complete reduction of ClO 4 and NO3 in S-compartment can stoichiometrically produce 2 254.2 mg/L SO2 production in the 4 . Actually, the average SO4 stage II–XI at steady state decreased from 270.2 ± 3.10 to 224.3 ± 2.28 mg/L with the decrease in HRT from 8 to 3 h. These results showed that S0 disproportionation in the continuous flow reactor could be related to HRT in S-compartment. The longer HRT could easily lead to S0 disproportionation due to the increase of the contact time between water and S0 particle. However, the contact chance of water and S0 particle decrease with the decrease in HRT, which can lead to the decrease of NO 3 and ClO4 removal and S0 disproportionation. Compared to the effluent from S-compartment, the SO2 concentration in the effluent from 4 H-compartment did not show obvious variation at different operational stages.
3.5. Variation of pH and ORP in the influent and effluent in different operational stages Fig. 7a shows pH variation in the influent and effluent from S-compartment and H-compartment in different operational stages. When the influent pH was between 7.25 and 7.40, the pH of effluent from S-compartment at steady state varied in the range
1013
of 6.95 and 7.15 in the operational stages I–XI. However, the pH in the effluent pH form H-compartment was between 7.20 and 7.30, which was always higher than the pH from S-compartment. According to Eqs. (1) and (2), the reduction processes of NO 3 and 0 ClO 4 using S as the electron donor simultaneously accompany with the generation of H+ and lead to low pH in the effluent from S-compartment. The H+ generated from S-compartment could be reduced as H2 by electrochemical process in H-compartment, and then the generated H2 as an electron donor was utilized to reduce NO 3 and ClO4 by hydrogen autotrophic reduction bacteria, which could explain that the pH in the effluent from H-compartment was always higher than that from S-compartment. Some researchers have reported that the effluent pH from sulfur-autotrophic denitrification and ClO 4 reduction was lower than the influent pH without the adjustment of alkaline matter [24,38]. Wang and Qu [37] also reported that the H+ generated from sulfur denitrification could be consumed by the bioelectrochemical hydrogen denitrification to achieve neutralization. Fig. 7b shows the variations of ORP in the effluents from S-compartment and H-compartment at different operational stages. As the influent was deoxygened in the influent tank, the ORP values in the influent varied in the range of 108 mV and 115 mV during the entire experiment. The ORP values in the effluent from S-compartment and H-compartment ranged from
Operational stage I
7.5
II
III
IV
V
a
VI
VII VIII
IX
X
XI
Influent Effluent from S-compartment Effluent from H-compartment
7.4
pH
7.3 7.2 7.1 7.0 6.9 0
20
40
60
80
100 120
140
160 180
200
220 240
Operational time (d) I
II
III
IV
V
VI
VII VIII
IX
X
XI
150 100
Influent Effluent from S-compartment Effluent from H-compartment
b
50
ORP mV
0 -50 -100
0
20
40
60
80
100 120
140 160
180 200
220
240
Operational time (d)
-150 -200 -250 -300 -350
Fig. 7. Variation of pH and ORP in the influent and effluent in different operational stages.
40 50 10 20 30 58 Distance from the bottom of the reactor (cm) Fig. 8. DGGE gel banding profiles of microbial communities along the flow path of the combined reactor.
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Table 3 Closest phylogenetic affiliations of sequences obtained from sludge samples along the flow path of the combined reactor. Bands
Closest related sequences
Accession number
Similarity (%)
Class containing related sequences
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35
Dechloromonas sp. JJ Aquabacterium commune (T) Uncultured bacterium SS-36 Uncultured Enterobacteriaceae bacterium Pseudomonas lundensis Uncultured bacterium p7n11ok Sulfurimonas autotrophica (T) Desulfocapsa thiozymogenes (T) Azonexus caeni (T) Sulfurimonas paralvinellae (T) Uncultured bacterium Thauera phenylacetica (T) Enterobacter sp. 2019 Uncultured bacterium Levilinea saccharolytica (T) Pseudorhodoferax soli (T) Methylibium petroleiphilum (T) Micropruina glycogenica (T) Dietzia natronolimnaea Sulfuricella denitrificans (T) Kineosphaera limosa (T) Hydrogenophaga sp. FS13–1 Acidovorax sp. BSB421 Paracoccus sp. ’CJSPY1 (P-I)’ Thioalkalivibrio thiocyanodenitrificans Uncultured bacterium Sulfuritalea hydrogenivorans (T) Thauera sp. R-28312 Zoogloea caeni (T) Enterobacter sp. LY402 Thiobacillus thioparus Amaricoccus kaplicensis (T) Azoarcus tolulyticus Thiobacillus denitrificans Staphylococcus sp. VM5
AY032611 AF035054 AY945866 EF562210 EU434368 FJ479276 AB088431 X95181 AB166882 AB252048 AB933384 AJ315678 JX566554 GU083529 AB109439 EU825700 AF176594 AB012607 AJ717372 AB506456 AF109792 JQ799975 Y18617 EF205262 AY360060 HM007537 AB552842 AM084110 DQ413148 DQ659161 HM535225 U88041 AF123075 AJ243144 DQ238834
97 99 100 100 99 99 91 99 90 92 99 95 96 100 92 92 93 98 100 91 96 92 99 95 92 100 93 93 94 100 99 98 93 97 99
b-proteobacteria b-proteobacteria – c-proteobacteria c-proteobacteria – e-proteobacteria d-proteobacteria b-proteobacteria e-proteobacteria – b-proteobacteria c-proteobacteria – Anaerolineaceae b-proteobacteria b-proteobacteria Actinobacteria Actinobacteria b-proteobacteria Actinobacteria b-proteobacteria b-proteobacteria a-proteobacteria c-proteobacteria – b-proteobacteria b-proteobacteria b-proteobacteria c-proteobacteria b-proteobacteria a-proteobacteria b-proteobacteria b-proteobacteria Bacilli
180 mV to 205 mV and from 255 mV to 250 mV from I to XI stage, respectively. The ORP values firstly illustrated some fluctuations after the decrease of HRT (or the increase of current intensity) and then trended to relatively stable ORP values. Suthersan [39] reported that the ORP values over 100 mV were interpreted to indicate an aerobic environment, whereas ones less than 100 mV were to indicate an anaerobic environment. The results in the present study showed that the ORP values in the effluent from both S-compartment and H-compartment were less than 180 mV, which illustrated good anaerobic conditions for the reduction of NO 3 and ClO4 in the combined reactor. 3.6. Microbial community analysis along the flow path in the combined bioreactor The microbial communities of ClO 4 and NO3 reduction on day 252 were investigated along the flow path of the combined reactor by DGGE. As shown in Fig. 8, the DGGR profile illustrated that some obvious variations were found in the band number and band intensity at different locations of the combined reactor. The prominent bands (i.e. bands 1, 9–11, 14, 21, and 29) were all found at the height of 10 cm, 20 cm, 30 cm, 40 cm, 50 cm and 58 cm from the bottom of the combined reactor, which were apparently stronger than the other bands. Some bands (i.e. bands 3–5, 12, 16, and 34) appeared or disappeared at certain locations in the combined reactor. The variations of band number and band intensity in the DGGE profiles might be related to the variation of ClO 4 and NO3 concentrations along the flow path of the combined reactor. To analyze further the microbial communities at different locations in the combined reactor, thirty-five visible bands from the DGGE profile were excised and then sequenced (Table 3). The
intensities of bands 1, 2, 8, 20, 29 and 34 decreased along the flow path of the combined reactor, suggesting the corresponding biomass tended to the descending trends. The bands 1 and 2 were most similar to Dechloromonas sp. and Aquabacterium commune, respectively. Some researchers reported that Dechloromonas sp. 0 could reduce both ClO 4 and NO3 by using S or H2 as electron donors [24,40–41]. Kalmbach et al. [41] found that A. commune sp. could utilize nitrate as an alternative electron acceptor, where they could use nitrite, chlorate, sulfate or iron (III) as electron acceptors. The bands 20 and 34 were close to Sulfuricella denitrificans and Thiobacillus denitrificans, respectively. Both S. denitrificans and T. denitrificans are anaerobic chemolithoautotrophs, which 0 could use NO 3 and nitrite (NO2 ) as electron acceptors and S as electron donor [42,43]. The sequence of band 8 matching Desulfocapsa thiozymogenes could grow by the disproportionation of S0 and by the disproportionation of sulfur oxyanions [44], which might lead the increase of SO2 4 concentration in the effluent. Azonexus caeni (band 9), Sulfurimonas paralvinellae (band 10) and Kineosphaera limosa (band 21) existed at different locations in the combined reactor. Thrash et al. [45] reported that A. caeni could utilize ClO 4 and NO3 as terminal electron acceptors and inorganic matters as electron donors. Takai et al. [46] illustrated that S. paralvinellae was a hydrogen- and sulfur-oxidizing chemolithoautotrophic denitrifying bacteria, suggesting that they might reduce 0 NO 3 or ClO4 by using S and H2 as electron donors in the combined reactor. The bands 28, 32 and 33 were related to Thauera sp. R28312, Amaricoccus kaplicensis and Azoarcus tolulyticus, respectively, which mainly appeared at the height of 40 cm and 56 cm from the bottom of the combined reactor. The genera of Thauera sp. and Azoarcus sp. were reported that they could reduce NO 3 and ClO 4 under autotrophic conditions [47,48]. The sequence of
M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016
band 31 matching to Thiobacillus thioparus was obligatory chemolithoautotrophic sulfur-oxidizing bacteria [49]. These results illustrates that some NO 3 reducer adapted to utilize ClO4 as an electron acceptor. 4. Conclusions The present showed that the simultaneous removal of perchlorate and nitrate could be achieved in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. The H+ generated from S-compartment could be consumed by electrochemical process in H-compartment. The removal efficiencies of NO 3 and ClO 4 in the S-compartment decreased with the increase of HRT, whereas they in the H-compartment were affected by current intensity. The sulfur- and hydrogen-autotrophic denitrifying process favored over the process of sulfur- and hydrogen-autotrophic perchlorate reduction in the combined reactor. The ORP values illustrated good anaerobic conditions for the reduction of NO 3 and ClO 4 in the combined reactor. Some sulfur- and hydrogen-autotrophic denitrifying bacteria could use both nitrate and ClO 4 as electron donors or only use NO 3 as an electron donor. Acknowledgement The work is founded by the National Natural Science Foundation of China (No. 21077096). References [1] Y. Cang, D.J. Roberts, D.A. Clifford, Development of cultures capable of reducing perchlorate and nitrate in high salt solutions, Water Res. 38 (2004) 3322–3330. [2] J.D. Coates, L.A. Achenbach, Microbial perchlorate reduction: rocket-fuelled metabolism, Nat. Rev. Microbiol. 2 (2004) 569–580. [3] B.E. Logan, Assessing the outlook for perchlorate remediation, Environ. Sci. Technol. 35 (2001) 482–487. [4] J. Wolff, Perchlorate and the thyroid gland, Pharmacol. Rev. 50 (1998) 89–105. [5] C.D. Rocca, V. Belgiorno, S. Meric, Overview of in-situ applicable nitrate removal processes, Desalination 204 (1–3) (2007) 46–62. [6] B. Gu, G.M. Brown, C.C. Chiang, Treatment of perchlorate-contaminated groundwater using highly selective, regenerable ion-exchange technologies, Environ. Sci. Technol. 41 (2007) 6277–6282. [7] L. Ye, S.T. Wang, H. You, J. Yao, X. Kang, Photocatalytic reduction of perchlorate in aqueous solutions in UV/Cu-TiO2/SiO2 system, Chem. Eng. J. 226 (2013) 434–443. [8] Y. Yoon, G. Amy, J. Cho, N. Her, J. Pellegrino, Transport of perchlorate (ClO 4) through NF and UF membranes, Desalination 147 (2002) 11–17. [9] Y.H. Xie, S.Y. Li, F. Wang, G.L. Liu, Removal of perchlorate from aqueous solution using protonated cross-linked chitosan, Chem. Eng. J. 156 (2010) 56– 63. [10] Y. Wang, L.Y. Jin, M.A. Deshusses, M.R. Matsumoto, The effects of various amendments on the biostimulation of perchlorate reduction in laboratory microcosm and flowthrough soil columns, Chem. Eng. J. 232 (2013) 388–396. [11] R. Epsztein, O. Nir, O. Lahav, M. Green, Selective nitrate removal from groundwater using a hybrid nanofiltration-reverse osmosis scheme, Chem. Eng. J. 279 (2015) 388–396. [12] W.R. Zhao, X. Zh., Y. Wang, Z.Y. Ai, D.Y. Zhao, Catalytic reduction of aqueous nitrates by metal supported catalysis on Al particles, Chem. Eng. J. 254 (2014) 410–417. [13] J.Y. Shen, Y. Chen, S.J. Wu, H.B. Wu, X.D. Liu, X.Y. Sun, J.S. Li, L.J. Wang, Enhanced pyridine biodegradation under anoxic condition: the key role of nitrate as the electron acceptor, Chem. Eng. J. 254 (2014) 410–417. [14] H. Choi, J. Silverstein, Inhibition of perchlorate reduction by nitrate in a fixed biofilm reactor, J. Hazard. Mater. 159 (2008) 440–445. [15] P.B. Hatzinger, Perchlorate biodegradation for water treatment, Environ. Sci. Technol. 39 (2005) 239–247. [16] Y. Xiao, D.J. Roberts, G. Zuo, M. Badruzzaman, G.S. Lehman, Characterization of microbial population in pilot-scale fluidized-bed reactors treating perchlorateand nitrate-laden brine, Water Res. 44 (2010) 4029–4036. [17] H. Zhang, B.E. Logan, J.M. Regan, L.A. Achenbach, M.A. Bruns, Molecular assessment of inoculated and indigenous bacteria in biofilms from a pilotscale perchlorate-ducing bioreactor, Microbial Ecol. 49 (2005) 388–398. [18] J.C. Thrash, J.I. Van Trump, K.A. Weber, E. Miller, L.A. Achenbach, J.D. Coates, Electrochemical stimulation of microbial perchlorate reduction, Environ. Sci. Technol. 41 (2007) 1740–1746. [19] B.E. Logan, D. LaPoint, Treatment of perchlorate- and nitrate-contaminated groundwater in an autotrophic, gas phase, packed-bed bioreactor, Water Res. 36 (2002) 3647–3653.
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Chemical Engineering Journal journal homepage: www.elsevier.com/locate/cej
Simultaneous removal of perchlorate and nitrate in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy Mengchun Gao a, Sen Wang b, Yun Ren b, Chunji Jin a, Zonglian She a,⇑, Yangguo Zhao a, Shiying Yang a, Liang Guo a, Jian Zhang b, Zhiwei Li b a b
Key Lab of Marine Environment and Ecology, Ministry of Education, Ocean University of China, Qingdao 266100, China College of Environmental Science and Engineering, Ocean University of China, Qingdao 266100, China
h i g h l i g h t s
0
S - and H2-autotrophy for NO3 and ClO4 reduction were investigated in a reactor. Autotrophic denitrifying process favored over autotrophic perchlorate reduction. +
0
H from S autotrophy process was consumed by electrochemical H2 autotrophy.
Some autotrophic denitrifier can use both NO3 and ClO4 as electron acceptor. Microbial communities at different positions in the reactor were analyzed.
a r t i c l e
i n f o
Article history: Received 23 June 2015 Received in revised form 24 September 2015 Accepted 25 September 2015
Keywords: Perchlorate Nitrate Sulfur autotrophy Electrochemical hydrogen autotrophy S0 particle
a b s t r a c t The removal of perchlorate (ClO 4 ) and nitrate (NO3 ) from drinking water was investigated in a combined reactor of sulfur autotrophy (S-compartment) and electrochemical hydrogen autotrophy (H-compartment). The removal efficiencies of NO and ClO in the S-compartment and 3 4 H-compartment were affected by hydraulic retention time (HRT) and current intensity, respectively. The sulfur- and hydrogen-autotrophic denitrifying process favored over the process of sulfur- and hydrogen-autotrophic perchlorate reduction in the combined reactor. The longer HRT could lead to sulfur (S0) disproportionation due to the increase of the contact time between water and S0 particle. The H+ generated from S-compartment could be reduced as H2 by electrochemical process in H-compartment, and the generated H2 as an electron donor was utilized to reduce NO 3 and ClO4 by hydrogen autotrophic reduction. The oxidation reduction potential (ORP) in the effluent from S-compartment and H-compartment were below 180 mV, suggesting good anaerobic conditions for the reduction of NO 3 and ClO 4 in the combined reactor. Some sulfur- and hydrogen-autotrophic denitrifying bacteria could use both NO3 and ClO4 as electron acceptors in the combined reactor. The DGGR profile illustrated that some variations were found in the microbial community at different locations of the combined reactor. Ó 2015 Elsevier B.V. All rights reserved.
1. Introduction Perchlorate (ClO 4 ) has been widely used in solid rocket fuels, missiles, explosives, pyrotechnics, and fireworks [1–3]. ClO 4 contamination in surface water and groundwater has become a significant environmental concern in recent years. ClO 4 can disrupt thyroid function by interfering with body’s intake of iodine, which inhibits the production of thyroid hormone [4]. Nitrate (NO 3 ) is ⇑ Corresponding author at: Key Lab of Marine Environment and Ecology, Ministry of Education, Ocean University of China, Qingdao 266100, China. Tel.: +86 532 66786351; fax: +86 532 66782810. E-mail address: [email protected] (Z. She). http://dx.doi.org/10.1016/j.cej.2015.09.082 1385-8947/Ó 2015 Elsevier B.V. All rights reserved.
often a co-contaminant with ClO 4 in surface water and groundwater, which mainly comes from agricultural runoff and wastewater discharge. The consumption of water contaminated by NO 3 can cause methemoglobinemia in infants, malformation and mutation [5]. The current technologies for ClO 4 and NO3 removal from drinking water and groundwater include ion exchange, photocatalytic reduction, membrane technology, absorption and biological reduction [6–13]. Ion exchange and biological reduction are regarded as the most cost–effective technologies for the treatment of ClO 4 and NO 3 contaminated water. The major disadvantage of ion exchange is that it can generate ClO 4 - and NO3 -laden brine, which requires specialized disposal and/or treatment. Biological reduction may
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M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016 completely transform ClO and N2, 4 and NO3 into innocuous Cl respectively. Some biological processes using different organic substrates as electron donors have been proved to be highly effec tive for ClO 4 and NO3 removal [14–17]. However, the residual organic substrate in the effluent can stimulate microbial growth in water distribution system and contribute to the formation of potentially toxic trihalomethanes during disinfection by chlorination [18]. To settle the above-mentioned problems, autotrophic reduction of ClO 4 and NO3 has been carried out by using inorganic matters as electron donors [19–21]. Hydrogen (H2) autotrophic reduction of ClO 4 and NO3 has been evaluated in some continuous flow bioreactors [19,22]. Although H2 is a good energy substrate for ClO 4 and NO3 reduction, it is difficult to handle in bulk quantities and is publicly perceived to represent a significant disaster threat due to its inherent explosive nature. Granular sulfur (S0) in packed bed bioreactors has been successfully used as an electron donor in the autotrophic reduction of ClO 4 and NO3 [20,23–26]. As granular S0 is insoluble in water, it can provide a slow-release supply of electron on demand and offer low expense cost as well as low maintenance cost. The stoichiometric equations for the S0 auto trophic reduction of ClO 4 and NO3 are respectively shown as follows [24,26]:
DC electrical source
+
_
Anode Carbon particle
Cathode
H2
10 cm
H-Part
H1 10 cm
S5 S4
þ
! 5:7 H þ
2:9 SO2 4
þ Cl þ 0:46 C5 H7 O2 N
ð1Þ
10 cm Sampling port 10 cm
S-Part
S3 10 cm
S2
2:9 S0 þ ClO4 þ 3:3 H2 O þ 1:8 CO2 þ 0:46 HCO3 þ 0:46 NHþ4
Effluent
S1
9 cm 1 cm S0 particle
1:1 S0 þ NO3 þ 0:76 H2 O þ 0:4 CO2 þ 0:08 NHþ4 ! 1:28 Hþ þ 1:1 SO2 4 þ 0:5 N2 þ 0:08 C5 H7 O2 N
Influent tank
ð2Þ
The main disadvantage for sulfur autotrophic ClO 4 and NO3 + reduction is the formation of sulfate (SO2 4 ) and H in the effluent. Limestone is often used as a low-cost alkalinity source, whereas it can increase the hardness of the effluent due to the release of Ca2+ during limestone dissolution. To make up for the deficiency of sulfur autotrophic ClO 4 and NO3 reduction, a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy was used to remove ClO 4 and NO3 in the present study. In the combined reactor, the H+ generated from sulfur-autotrophy ClO 4 and NO 3 reduction could be consumed by the bioelectrochemical hydrogen-autotrophy ClO and NO reduction to achieve 4 3 neutralization, and the SO2 concentration in the effluent can be 4 controlled by adjusting the ClO 4 and NO3 loading of sulfur autotrophy and electrochemical hydrogen autotrophy. The main objectives of the present study were (1) to evaluate the feasibility of ClO 4 reduction by sulfur- and hydrogenautotrophic denitrifying bacteria; (2) to investigate the effect of hydraulic retention time (HRT) and current intensity on ClO 4 and NO 3 reduction in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy; (3) to analyze the variation of microbial community along the flow path in the combined reactor.
stainless column. The space between the cathode and the anode in the H-compartment was filled with carbon particle with 3.0– 4.0 mm in diameter. Before the synthetic wastewater was fed to the combined reactor, it was firstly deoxidized by injecting nitrogen gas in the influent tank. A peristaltic pump was used to feed the influent into the combined reactor, and direct current was supplied to the H-compartment by a silicon rectifier. The Chemical processes of NO 3 and ClO4 reduction in the S-compartment and H-compartment is shown in Fig. 2. The water sampling ports were mounted at the height of 1 cm (S1), 10 cm (S2), 20 cm (S3), 30 cm (S4), 40 cm (S5), 50 cm (H1) and 60 cm (H2) from the bottom of the combined reactor. In addition, the S0 granules in the S-compartment were not replenished during the whole operational periods.
2. Materials and methods
2.2. Inoculated sludge and synthetic wastewater
2.1. Experimental set-up
The inoculated sludge was taken from an anaerobic tank of Lichunhe wastewater treatment plant in Qingdao city, China. The synthetic wastewater fed to the combined reactor was prepared daily by adding NaNO3, NaClO4H2O, and nutrient medium to the tap water. The compositions of nutrient medium were shown as follows: NH4HCO3 0.410 g/L, NaHCO3 2.700 g/L, MgSO47H2O 0.100 g/L, Ca(OH)2 0.005 g/L, and trace element solution 1 mL/L. The trace element solution consisted of the following components (g/L): Na2SeO35H2O 0.1, H3BO3 0.05, MnSO4H2O 0.415, NiSO46H2O 0.113, FeSO47H2O 2.8, Na2WO42H2O 0.5, CoSO412H2O 2.36, KAl (SO4)212H2O 0.175, EDTA 1.0, ZnSO47H2O 0.106, (NH4)6Mo7O24H2O 0.05, and CuSO45H2O 0.157.
The schematic diagram of an experimental set-up is shown in Fig. 1, which consists of sulfur autotrophy (S-compartment) and electrochemical hydrogen autotrophy (H-compartment). The Scompartment was a cylindrical plexiglass column with 7.5 cm in diameter and 40 cm in height, which was filled with S0 granules of 3.0–4.0 mm in diameter. A cylindrical stainless column with 7.5 cm in diameter and 25 cm in height was used as the cathode in the H-compartment. A graphite rod with 2.5 cm in diameter and 25 cm in height was used as the anode in the Hcompartment, which was located in the center of the cylindrical
Influent pump
Fig. 1. Schematic diagram of an experimental set-up.
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M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016
2H2O +2e
ClO4NO3-
Electrolysis
2OH+ Electrochemical reduction H2 H2O 20 SO4 + 2e S -2e-6eClO4 Cl- + N2 Cl- + N2 + 2H+ NO3- H2 autotrophic bacteria S0 autotrophic bacteria S - c o m p a r t m e nt
H-compartment
Fig. 2. Chemical processes of NO 3 and ClO4 reduction in the S-compartment and H-compartment.
Table 1 Operational conditions of reactor #1. Operational stages Operational time (d)
I 0–20
II 20–32
III 32–50
IV 50–64
V 64–84
VI 84–104
VII 104–122
VIII 122–168
Influent NO 3 -N concentration (mg/L) Influent ClO 4 concentration (mg/L) HRT (h) Current intensity (mA) Voltage (V)
28 0 12 10 2
28 0 8 10 2
28 0 4 10 2
28 0 2 10 2
28 0 1 10 2
28 0 0.5 10 2
28 0 0.5 20 4
0 124.38 0.5 20 4
Table 2 Operational conditions of reactor 2#. Operational periods Operational days (d)
I 0–38
II 38–58
III 58–80
IV 80–106
V 106–126
VI 126–152
VII 152–170
VIII 170–188
IX 188–214
X 214–238
XI 238–252
Influent ClO 4 concentration (mg/L) Influent NO 3 -N concentration (mg/L) HRT (h) Current intensity (mA) Voltage (V)
2 20 12 10 2
10 3 8 10 2
10 30 5 10 2
10 30 3 10 2
10 30 2 10 2
10 30 1 10 2
10 30 0.5 10 2
10 30 0.5 15 3
10 30 0.5 25 5
10 30 0.5 35 7
10 30 0.5 45 9
2.3. Operational procedure
3. Results and discussion
Two identical experimental set-ups were prepared in the present study. The first reactor (#1) was used to evaluate the probability of ClO 4 reduction by sulfur- and hydrogen-autotrophic denitrifying bacteria. The second reactor (#2) was used to investi gate the effect of HRT and current intensity on ClO 4 and NO3 reduction in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. The operational procedures of reactor #1 and #2 were summarized in Tables 1 and 2, respectively.
3.1. Feasibility assessment of perchlorate reduction by autotrophic denitrifying bacteria in the combined reactor
2.4. Analytical methods The water samples were filtered using a 0.22 lm filter membrane and then stored in a refrigerator at 4 °C for further analysis. The oxidation reduction potential (ORP) was measured by a portable ORT meter. A portable pH meter (Model 6010, WTW, Germany) was used to monitor pH in the influent and effluent. NO 3 , ClO4 , and SO2 were determined by an ion chromatography (ICS-300, 4 Dionex, USA). In order to evaluate the distribution of microbial community along the flow path of reactor #2, the biofilm samples were obtained at the height of 10 cm, 20 cm, 30 cm, 40 cm, 50 cm and 58 cm from the bottom at the end of the experiment (day 252), respectively. The DNA extraction, denaturing gradient gel electrophoresis (DGGE) analysis and sequencing were performed according to the methodology reported by Wang et al. [27].
Most perchlorate-reducing bacteria can utilize NO 3 as an alternative electron acceptor under anoxic conditions [2,28]. Some denitrifying bacteria can reduce ClO 4 using nitrate reductase enzymes as well as specialized perchlorate reductases [29,30]. However, some researchers have also found that the common denitrifying bacteria cannot use chlorate (ClO 3 ) or ClO4 as an electron acceptor [28,31–33]. In order to valuate the feasibility of ClO 4 reduction by autotrophic denitrifying bacteria, the ClO 4 reduction by sulfur- and hydrogen-autotrophic denitrifying bacteria was investigated in the combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy at different conditions (Table 1). According to Eqs. (1) and (2), the complete reductions of NO 3 and ClO4 per mol need to transfer 5 mol e and 8 mol e, respectively. Considering the equilibrium of electron transfer, the influent concentrations of NO 3 -N in stages I–VII and ClO4 in stage VIII were 28 and 124.38 mg/L, respectively, corresponding to 2 mmol/L NO 3 and 1.25 nmol/L ClO 4 in the influent. Fig. 3 shows the variation of NO (or ClO concentration in the effluent from 3 -N 4) S-compartment and H-compartment in different operational stages. In the operational stages I–V, the NO 3 -N in the effluent from both S-compartment and H-compartment at steady state was nearly not detected under the conditions of HRT varying from 12 to 1 h and 10 mA current. When the HRT was shortened to 0.5 h in the operational stage VI, the average NO 3 -N concentrations in
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Operational stage
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Effluent NO 3--N from H-com
Inffluent ClO 4-
Effluent ClO 4- from S-com
Effluent ClO 4- from H-com
10 9
2.0
ClO4- concentration (mg/L)
ClO4- or NO3--N concentration (mmol/L)
2.5
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1.0
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8
Effluent ClO 4- from S-com
7
Effluent ClO 4- from H-com
6 5 4 3 2
0.5
1 0 0.0
0 0
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40
60 80 100 Operational time (d)
120
140
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40
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80
160
100 120
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160 180
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220 240
IX
X
Operational time (d) I
Fig. 3. Nitration reduction (or perchlorate reduction) by autotrophic denitrifying bacteria in the combined reactor. S-compartment, S-com; H-Compartment, H-com.
II
III
IV
V
VI
VII VIII
XI
35
3.2. Simultaneous removal of ClO 4 and NO3 in different operational stages
Some changes were made for the combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. These changes included the increase in current intensity from 10 to 45 mA and the decrease in HRT from 12 to 0.5 h (Table 2). The NO 3 and ClO4 concentrations in the effluent were consistent for at least 10 days in an operational stage before the next operational stage began in the combined reactor. Fig. 4 shows the removal effi ciencies of ClO 4 and NO3 in the combined reactor in different operational stages. In the operational stage I, the reactor was fed 20 mg/L NO 3 -N and 2 mg/L ClO4 under the conditions of 12 h HRT and 10 mA current. No NO in the effluent from 3 S-compartment was detected at steady state, suggesting the H-compartment did not participate in the process of NO 3 reduction. The average ClO 4 concentration in the effluent from S-compartment at steady states was 0.49 ± 0.01 mg/L in the operational stage I, whereas no ClO 4 was detected in the effluent from H-compartment. In the operational stages II–VI, the reactor was fed 30 mg/L NO 3 -N and 10 mg/L ClO4 under the conditions of 10 mA current and HRT varying from 8 to 1 h. The effluent NO 3 from the S-compartment at steady state was not detected in operational stage II–VI, which illustrated that the influent NO 3 could be
-
NO3 -N concentration (mg/L)
30
the effluent from S-compartment and H-compartment at steady states were 8.86 ± 0.03 mg/L and 2.40 ± 0.01 mg/L at 10 mA current, respectively. In order to increase the denitrification efficiency of H-compartment, the current was increased to 20 mA for producing more H2 in the stage VII. As shown in Fig. 3, the average NO 3 -N concentrations in the effluent from S-compartment and H-compartment at steady states was 5.87 ± 0.02 mg/L and 0 mg/L in the operational stage VII, respectively. In the stage VIII, the concentrations of ClO 4 and NO3 in the influent were 124.38 and 0 mg/L, respectively. The average ClO 4 concentration in the effluent from S-compartment and H-compartment at steady states was 98.63 ± 0.22 mg/L and 94.53 ± 0.31 mg/L in the stage VIII, respectively. Compared with the NO 3 reduction in stage VII, the sulfur- and hydrogen-autotrophic denitrifying bacteria could partially reduce ClO 4 in the present study, suggesting some auto trophic denitrifying bacteria could use both NO 3 and ClO4 as electron acceptors or only use NO 3 as an electron acceptor.
Influent NO 3--N
25
Effluent NO 3--N from S-com
20
Effluent NO 3--N from H-com
15 10 5 0 0
20
40
60
80
100
120 140
160 180
200 220
240
Operational time (d) Fig. 4. Removal efficiencies of ClO 4 and NO3 -N in the combined reactor in different operational stages. S-compartment, S-com; H-Compartment, H-com.
removed in the S-compartment at 1–8 h HRT. The average effluent ClO 4 concentrations from S-compartment at steady states were 2.03 ± 0.06, 1.06 ± 0.02, 1.00 ± 0.01, 1.51 ± 0.03 and 2.04 ± 0.05 mg/L at the HRT of 8, 5, 3, 2 and 1 h, respectively. However, no ClO 4 in the effluent from H-compartment at steady states was detected in operational stage II–VI. In the operational stage VII, the average NO 3 -N and ClO4 concentrations in the effluent from S-compartment increased to 8.92 ± 0.06 and 4.98 ± 0.04 mg/L as the HRT decreased to 0.5 h, respectively. Similarly, the average NO and ClO concentrations in the effluent from 3 -N 4 H-compartment increased to 2.48 ± 0.03 and 3.99 ± 0.10 mg/L in the operational stage VII, respectively. Compared to the stage VI, the NO concentrations in the effluent from 3 -N and ClO4 H-compartment increased in the stage VII due to the shortage of HRT, which was peculated that H2 as the electron donor in H2-compartment could be inadequate for the reduction of NO-N and ClO 4 in stage VII. In order to enhance the hydrogen autotrophic reduction of NO 3 and ClO4 in the H-compartment, the current intensity increased from 10 to 45 mA with the conversion of the operational stage from V II to XI. Compared with the operational stage VII, the average effluent NO 3 and ClO4 concentrations from S-compartment had no obvious variation, suggesting the removal efficiencies of NO 3 and ClO4 were related to HRT in the S-compartment. However, No NO was detected from 3 H-compartment in the operational stages VII–XI, and the average effluent ClO 4 from H-compartment decreased from 3.99 ± 0.10 to
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1.49 ± 0.08 mg/L with the increase in current from 10 to 45 mA. As H2 as the electron donor in the H-compartment originated from the electrochemical reduction of H+ from the S-compartment or water electrolysis process, the H2 production was closely related to the current intensity in the H2-compartment. The present results suggested that the removal efficiencies of NO 3 and ClO4 in the H-compartment could be affected by the variation of current intensity. 3.3. Variation of ClO 4 and NO3 concentrations along the flow path of the combined reactor In order to investigate the relative progress of ClO 4 and NO3 reduction in the combined reactor (#2), the variations of ClO 4 and NO 3 concentrations at steady state were analyzed along the flow path of the combined reactor. The concentrations of ClO 4 and NO 3 decreased along the flow path of the combined reactor in different operational stages. This phenomenon could be explained that sulfur- (or hydrogen-) autotrophic reduction bacteria and denitrifying bacteria coexisted in S-compartment (or H-compartment) or that some autotrophic bacteria could simulta neously utilize NO 3 and ClO4 as electron acceptors. As shown in Fig. 5, no NO3 was detected in the S4 sampling port under the conditions of 10 mA current and HRT varying from 3 to 1 h, whereas the effluent ClO 4 concentration from the S4 sampling port increased from 4 to 6 mg/L with the decrease in HRT from 3 to 1 h. No ClO 4 in the effluent was found in the sampling port of H1, H2 and H2 at HRT of 3, 2 and 1 h, respectively. The NO 3 -N and ClO 4 concentrations from the sampling port H2 were 3 and
HRT =3h, I=10mA HRT =2h, I=10mA HRT =1h, I=10mA HRT =0.5h, I=10mA
ClO4- concentration (mg/L)
10
8
HRT =0.5h, I=15mA HRT =0.5h, I=25mA HRT =0.5h, I=35mA
6
4
2
5 mg/L under 10 mA current intensity and 0.5 h HRT, respectively. The ClO 4 concentration from sampling port H2 decreased with the increase in current from 10 to 35 mA, whereas no NO 3 from sampling port H1, H1 and H2 was detected under the conditions of 15–35 mA current intensity and 0.5 h HRT. The abovementioned results showed that the processes of autotrophic perchlorate reduction and denitrification could occur in parallel when sufficient substrates were present in the combined reactor. Com pared to NO 3 reduction, a lag in ClO4 reduction was found in the S-compartment and H-compartment of the combined reactor. According to the reports of Ontiveros-Valencia et al. [34,35], the present results could be explained that some microorganisms pre fer NO 3 over ClO4 as electron acceptor, which delayed the onset of sulfur- and hydrogen-autotrophic ClO 4 reduction. The sulfur- (or hydrogen-) autotrophic ClO 4 reduction might begin and occur simultaneously with the sulfur- (or hydrogen-) autotrophic denitrification before NO 3 was completely removed from the influent containing NO 3 and ClO4 . In other words, some sulfur- (or hydrogen-) autotrophic denitrifying bacteria in the present study might not utilize ClO 4 as an electron acceptor, and others could remove both ClO 4 and NO3 or retain the capacity for ClO4 reduc tion when exposed to both ClO4 and NO3 . Ju et al. [20] reported 0 that the addition of NO 3 to the S -oxidizing perchlorate reduction bacteria delay the onset of ClO 4 reduction until NO3 was partially consumed. 3.4. Variation of SO2 4 concentration in the effluent from the combined reactor 0 The coupling of ClO 4 and NO3 reduction with S oxidation can concomitantly produce SO2 in S-compartment. According to Eqs. 4 (1) and (2), the SO2 4 production from ClO4 and NO3 reduction in S-compartment is stoichiometrically 2.80 mg SO2 4 /mg ClO4 and 7.54 mg SO2 /mg NO -N, respectively. As shown in Fig. 6, the aver4 3 age effluent SO2 concentration at steady states was 4 265.24 ± 4.84 mg/L from S-compartment when the influent with 20 mg/L NO 3 -N and 2 mg/L ClO4 was fed to the combined reactor at 12 h HRT in the operational stage I. The SO2 4 production in the operational stage I was much higher than the theoretical SO2 4 pro duction (156.4 mg/L) from NO 3 and ClO4 reduction. The production of excessive SO2 4 in the operational stage I showed that the sulfur disproportionation could happen in the reactor as follows:
þ 4S0 þ 4H2 O ! 3H2 S þ SO2 4 þ 2H
ð3Þ
0 S1
S2
S3
S4
S5
H1
H2
Operational stage
Sampling port
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30
HRT =3h, I=10mA HRT =2h, I=10mA HRT =1h, I=10mA HRT =0.5h, I=10mA HRT =0.5h, I=15mA HRT =0.5h, I=25mA HRT =0.5h, I=35mA
25 20
V
VI
VII VIII
IX
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Influent Effluent from S-compartment Effluent from H-compartment
260 240 SO42- concentration (mg/L)
NO3 --N concentration (mg/L)
II
15 10
220 200 180 160 140 120
5
100
0 S1
80
S2
S3
S4
S5
H1
H2
Sampling port Fig. 5. Variation of ClO 4 and NO3 -N concentrations along the flow path in the combined reactor.
0
20
40
60
80
100 120 140 160 180 Operational time (d)
200 220
240
Fig. 6. Variation of SO2 4 concentration in the effluent from the combined reactor in different operational stages.
M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016 Some researchers have reported that ClO 4 and NO3 reduction 0 0 using S as the electron donor coupled with S disproportionation in batch experiment or continuous flow experiment [24,36–37]. In the operational stages II–XI, the reactor was fed 30 mg/L NO 3 -N and 10 mg/L ClO 4 in the influent, and the complete reduction of ClO 4 and NO3 in S-compartment can stoichiometrically produce 2 254.2 mg/L SO2 production in the 4 . Actually, the average SO4 stage II–XI at steady state decreased from 270.2 ± 3.10 to 224.3 ± 2.28 mg/L with the decrease in HRT from 8 to 3 h. These results showed that S0 disproportionation in the continuous flow reactor could be related to HRT in S-compartment. The longer HRT could easily lead to S0 disproportionation due to the increase of the contact time between water and S0 particle. However, the contact chance of water and S0 particle decrease with the decrease in HRT, which can lead to the decrease of NO 3 and ClO4 removal and S0 disproportionation. Compared to the effluent from S-compartment, the SO2 concentration in the effluent from 4 H-compartment did not show obvious variation at different operational stages.
3.5. Variation of pH and ORP in the influent and effluent in different operational stages Fig. 7a shows pH variation in the influent and effluent from S-compartment and H-compartment in different operational stages. When the influent pH was between 7.25 and 7.40, the pH of effluent from S-compartment at steady state varied in the range
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of 6.95 and 7.15 in the operational stages I–XI. However, the pH in the effluent pH form H-compartment was between 7.20 and 7.30, which was always higher than the pH from S-compartment. According to Eqs. (1) and (2), the reduction processes of NO 3 and 0 ClO 4 using S as the electron donor simultaneously accompany with the generation of H+ and lead to low pH in the effluent from S-compartment. The H+ generated from S-compartment could be reduced as H2 by electrochemical process in H-compartment, and then the generated H2 as an electron donor was utilized to reduce NO 3 and ClO4 by hydrogen autotrophic reduction bacteria, which could explain that the pH in the effluent from H-compartment was always higher than that from S-compartment. Some researchers have reported that the effluent pH from sulfur-autotrophic denitrification and ClO 4 reduction was lower than the influent pH without the adjustment of alkaline matter [24,38]. Wang and Qu [37] also reported that the H+ generated from sulfur denitrification could be consumed by the bioelectrochemical hydrogen denitrification to achieve neutralization. Fig. 7b shows the variations of ORP in the effluents from S-compartment and H-compartment at different operational stages. As the influent was deoxygened in the influent tank, the ORP values in the influent varied in the range of 108 mV and 115 mV during the entire experiment. The ORP values in the effluent from S-compartment and H-compartment ranged from
Operational stage I
7.5
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a
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7.4
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7.3 7.2 7.1 7.0 6.9 0
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160 180
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b
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0 -50 -100
0
20
40
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80
100 120
140 160
180 200
220
240
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-150 -200 -250 -300 -350
Fig. 7. Variation of pH and ORP in the influent and effluent in different operational stages.
40 50 10 20 30 58 Distance from the bottom of the reactor (cm) Fig. 8. DGGE gel banding profiles of microbial communities along the flow path of the combined reactor.
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Table 3 Closest phylogenetic affiliations of sequences obtained from sludge samples along the flow path of the combined reactor. Bands
Closest related sequences
Accession number
Similarity (%)
Class containing related sequences
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35
Dechloromonas sp. JJ Aquabacterium commune (T) Uncultured bacterium SS-36 Uncultured Enterobacteriaceae bacterium Pseudomonas lundensis Uncultured bacterium p7n11ok Sulfurimonas autotrophica (T) Desulfocapsa thiozymogenes (T) Azonexus caeni (T) Sulfurimonas paralvinellae (T) Uncultured bacterium Thauera phenylacetica (T) Enterobacter sp. 2019 Uncultured bacterium Levilinea saccharolytica (T) Pseudorhodoferax soli (T) Methylibium petroleiphilum (T) Micropruina glycogenica (T) Dietzia natronolimnaea Sulfuricella denitrificans (T) Kineosphaera limosa (T) Hydrogenophaga sp. FS13–1 Acidovorax sp. BSB421 Paracoccus sp. ’CJSPY1 (P-I)’ Thioalkalivibrio thiocyanodenitrificans Uncultured bacterium Sulfuritalea hydrogenivorans (T) Thauera sp. R-28312 Zoogloea caeni (T) Enterobacter sp. LY402 Thiobacillus thioparus Amaricoccus kaplicensis (T) Azoarcus tolulyticus Thiobacillus denitrificans Staphylococcus sp. VM5
AY032611 AF035054 AY945866 EF562210 EU434368 FJ479276 AB088431 X95181 AB166882 AB252048 AB933384 AJ315678 JX566554 GU083529 AB109439 EU825700 AF176594 AB012607 AJ717372 AB506456 AF109792 JQ799975 Y18617 EF205262 AY360060 HM007537 AB552842 AM084110 DQ413148 DQ659161 HM535225 U88041 AF123075 AJ243144 DQ238834
97 99 100 100 99 99 91 99 90 92 99 95 96 100 92 92 93 98 100 91 96 92 99 95 92 100 93 93 94 100 99 98 93 97 99
b-proteobacteria b-proteobacteria – c-proteobacteria c-proteobacteria – e-proteobacteria d-proteobacteria b-proteobacteria e-proteobacteria – b-proteobacteria c-proteobacteria – Anaerolineaceae b-proteobacteria b-proteobacteria Actinobacteria Actinobacteria b-proteobacteria Actinobacteria b-proteobacteria b-proteobacteria a-proteobacteria c-proteobacteria – b-proteobacteria b-proteobacteria b-proteobacteria c-proteobacteria b-proteobacteria a-proteobacteria b-proteobacteria b-proteobacteria Bacilli
180 mV to 205 mV and from 255 mV to 250 mV from I to XI stage, respectively. The ORP values firstly illustrated some fluctuations after the decrease of HRT (or the increase of current intensity) and then trended to relatively stable ORP values. Suthersan [39] reported that the ORP values over 100 mV were interpreted to indicate an aerobic environment, whereas ones less than 100 mV were to indicate an anaerobic environment. The results in the present study showed that the ORP values in the effluent from both S-compartment and H-compartment were less than 180 mV, which illustrated good anaerobic conditions for the reduction of NO 3 and ClO4 in the combined reactor. 3.6. Microbial community analysis along the flow path in the combined bioreactor The microbial communities of ClO 4 and NO3 reduction on day 252 were investigated along the flow path of the combined reactor by DGGE. As shown in Fig. 8, the DGGR profile illustrated that some obvious variations were found in the band number and band intensity at different locations of the combined reactor. The prominent bands (i.e. bands 1, 9–11, 14, 21, and 29) were all found at the height of 10 cm, 20 cm, 30 cm, 40 cm, 50 cm and 58 cm from the bottom of the combined reactor, which were apparently stronger than the other bands. Some bands (i.e. bands 3–5, 12, 16, and 34) appeared or disappeared at certain locations in the combined reactor. The variations of band number and band intensity in the DGGE profiles might be related to the variation of ClO 4 and NO3 concentrations along the flow path of the combined reactor. To analyze further the microbial communities at different locations in the combined reactor, thirty-five visible bands from the DGGE profile were excised and then sequenced (Table 3). The
intensities of bands 1, 2, 8, 20, 29 and 34 decreased along the flow path of the combined reactor, suggesting the corresponding biomass tended to the descending trends. The bands 1 and 2 were most similar to Dechloromonas sp. and Aquabacterium commune, respectively. Some researchers reported that Dechloromonas sp. 0 could reduce both ClO 4 and NO3 by using S or H2 as electron donors [24,40–41]. Kalmbach et al. [41] found that A. commune sp. could utilize nitrate as an alternative electron acceptor, where they could use nitrite, chlorate, sulfate or iron (III) as electron acceptors. The bands 20 and 34 were close to Sulfuricella denitrificans and Thiobacillus denitrificans, respectively. Both S. denitrificans and T. denitrificans are anaerobic chemolithoautotrophs, which 0 could use NO 3 and nitrite (NO2 ) as electron acceptors and S as electron donor [42,43]. The sequence of band 8 matching Desulfocapsa thiozymogenes could grow by the disproportionation of S0 and by the disproportionation of sulfur oxyanions [44], which might lead the increase of SO2 4 concentration in the effluent. Azonexus caeni (band 9), Sulfurimonas paralvinellae (band 10) and Kineosphaera limosa (band 21) existed at different locations in the combined reactor. Thrash et al. [45] reported that A. caeni could utilize ClO 4 and NO3 as terminal electron acceptors and inorganic matters as electron donors. Takai et al. [46] illustrated that S. paralvinellae was a hydrogen- and sulfur-oxidizing chemolithoautotrophic denitrifying bacteria, suggesting that they might reduce 0 NO 3 or ClO4 by using S and H2 as electron donors in the combined reactor. The bands 28, 32 and 33 were related to Thauera sp. R28312, Amaricoccus kaplicensis and Azoarcus tolulyticus, respectively, which mainly appeared at the height of 40 cm and 56 cm from the bottom of the combined reactor. The genera of Thauera sp. and Azoarcus sp. were reported that they could reduce NO 3 and ClO 4 under autotrophic conditions [47,48]. The sequence of
M. Gao et al. / Chemical Engineering Journal 284 (2016) 1008–1016
band 31 matching to Thiobacillus thioparus was obligatory chemolithoautotrophic sulfur-oxidizing bacteria [49]. These results illustrates that some NO 3 reducer adapted to utilize ClO4 as an electron acceptor. 4. Conclusions The present showed that the simultaneous removal of perchlorate and nitrate could be achieved in a combined reactor of sulfur autotrophy and electrochemical hydrogen autotrophy. The H+ generated from S-compartment could be consumed by electrochemical process in H-compartment. The removal efficiencies of NO 3 and ClO 4 in the S-compartment decreased with the increase of HRT, whereas they in the H-compartment were affected by current intensity. The sulfur- and hydrogen-autotrophic denitrifying process favored over the process of sulfur- and hydrogen-autotrophic perchlorate reduction in the combined reactor. The ORP values illustrated good anaerobic conditions for the reduction of NO 3 and ClO 4 in the combined reactor. Some sulfur- and hydrogen-autotrophic denitrifying bacteria could use both nitrate and ClO 4 as electron donors or only use NO 3 as an electron donor. Acknowledgement The work is founded by the National Natural Science Foundation of China (No. 21077096). References [1] Y. Cang, D.J. Roberts, D.A. Clifford, Development of cultures capable of reducing perchlorate and nitrate in high salt solutions, Water Res. 38 (2004) 3322–3330. [2] J.D. Coates, L.A. Achenbach, Microbial perchlorate reduction: rocket-fuelled metabolism, Nat. Rev. Microbiol. 2 (2004) 569–580. [3] B.E. Logan, Assessing the outlook for perchlorate remediation, Environ. Sci. Technol. 35 (2001) 482–487. [4] J. Wolff, Perchlorate and the thyroid gland, Pharmacol. Rev. 50 (1998) 89–105. [5] C.D. Rocca, V. Belgiorno, S. Meric, Overview of in-situ applicable nitrate removal processes, Desalination 204 (1–3) (2007) 46–62. [6] B. Gu, G.M. Brown, C.C. Chiang, Treatment of perchlorate-contaminated groundwater using highly selective, regenerable ion-exchange technologies, Environ. Sci. Technol. 41 (2007) 6277–6282. [7] L. Ye, S.T. Wang, H. You, J. Yao, X. Kang, Photocatalytic reduction of perchlorate in aqueous solutions in UV/Cu-TiO2/SiO2 system, Chem. Eng. J. 226 (2013) 434–443. [8] Y. Yoon, G. Amy, J. Cho, N. Her, J. Pellegrino, Transport of perchlorate (ClO 4) through NF and UF membranes, Desalination 147 (2002) 11–17. [9] Y.H. Xie, S.Y. Li, F. Wang, G.L. Liu, Removal of perchlorate from aqueous solution using protonated cross-linked chitosan, Chem. Eng. J. 156 (2010) 56– 63. [10] Y. Wang, L.Y. Jin, M.A. Deshusses, M.R. Matsumoto, The effects of various amendments on the biostimulation of perchlorate reduction in laboratory microcosm and flowthrough soil columns, Chem. Eng. J. 232 (2013) 388–396. [11] R. Epsztein, O. Nir, O. Lahav, M. Green, Selective nitrate removal from groundwater using a hybrid nanofiltration-reverse osmosis scheme, Chem. Eng. J. 279 (2015) 388–396. [12] W.R. Zhao, X. Zh., Y. Wang, Z.Y. Ai, D.Y. Zhao, Catalytic reduction of aqueous nitrates by metal supported catalysis on Al particles, Chem. Eng. J. 254 (2014) 410–417. [13] J.Y. Shen, Y. Chen, S.J. Wu, H.B. Wu, X.D. Liu, X.Y. Sun, J.S. Li, L.J. Wang, Enhanced pyridine biodegradation under anoxic condition: the key role of nitrate as the electron acceptor, Chem. Eng. J. 254 (2014) 410–417. [14] H. Choi, J. Silverstein, Inhibition of perchlorate reduction by nitrate in a fixed biofilm reactor, J. Hazard. Mater. 159 (2008) 440–445. [15] P.B. Hatzinger, Perchlorate biodegradation for water treatment, Environ. Sci. Technol. 39 (2005) 239–247. [16] Y. Xiao, D.J. Roberts, G. Zuo, M. Badruzzaman, G.S. Lehman, Characterization of microbial population in pilot-scale fluidized-bed reactors treating perchlorateand nitrate-laden brine, Water Res. 44 (2010) 4029–4036. [17] H. Zhang, B.E. Logan, J.M. Regan, L.A. Achenbach, M.A. Bruns, Molecular assessment of inoculated and indigenous bacteria in biofilms from a pilotscale perchlorate-ducing bioreactor, Microbial Ecol. 49 (2005) 388–398. [18] J.C. Thrash, J.I. Van Trump, K.A. Weber, E. Miller, L.A. Achenbach, J.D. Coates, Electrochemical stimulation of microbial perchlorate reduction, Environ. Sci. Technol. 41 (2007) 1740–1746. [19] B.E. Logan, D. LaPoint, Treatment of perchlorate- and nitrate-contaminated groundwater in an autotrophic, gas phase, packed-bed bioreactor, Water Res. 36 (2002) 3647–3653.
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