Improved anammox performance with a flow switched anaerobic baffled reactor (FSABR) modified from a common anaerobic baffled reactor (CABR)

Ecological Engineering 92 (2016) 229–235

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Improved anammox performance with a flow switched anaerobic baffled reactor (FSABR) modified from a common anaerobic baffled reactor (CABR) Hui Chen a,b , Qian-Qian Chen a,b , Zhi-Jian Shi a,b , Jia-Jia Xu a,b , An-Na Liu a,b , Li-Ling He a,b , Yu-Huan Wu a,b , Man-Ling Shi a,b , Ren-Cun Jin a,b,∗ a b

College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou310036, China Key Laboratory of Hangzhou City for Ecosystem Protection and Restoration, Hangzhou Normal University, Hangzhou 310036, China

a r t i c l e

i n f o

Article history: Received 23 May 2015 Received in revised form 11 April 2016 Accepted 15 April 2016 Available online 28 April 2016 Keywords: Anammox ABR Feeding regime Sludge characteristics

a b s t r a c t An anaerobic ammonium oxidation (anammox) reactor was operated with two feeding regimes. In stage 1, the reactor was operated as a common anaerobic baffled reactor (CABR) (constant flow direction), whereas in stage 2, the reactor was modified as a flow switched anaerobic baffled reactor (FSABR) with the flow direction switching every 10 days. Adjusting the feeding regime resulted in an increase in the average nitrogen loading rate (from 1.62 to 1.80 kgN m−3 d−1 ) and nitrogen removal efficiency (from 75.4 to 83.1%). In addition, a maximum nitrogen removal rate of 3.49 kgN m−3 d−1 was obtained in stage 2, compared with 3.00 kgN m−3 d−1 in stage 1. The sludge properties were also enhanced; from stage 1 to stage 2, the specific anammox activity increased by more than 5-fold, the settling velocity increased from 64.1 to 71.9 m h−1 , and the average particle diameter increased from 0.8 to 2.3 mm. However, there was minimal variation in the heme c content. The maximum substrate removal rates in stages 1 and 2 were 4.28 and 46.38 kgN m−3 d−1 when fitted by the Stover-Kincannon model, respectively. The results indicate that the FSABR performance was significantly enhanced compared with the CABR. © 2016 Elsevier B.V. All rights reserved.

1. Introduction The anaerobic treatment of wastewater has been considered a promising process over aerobic treatment due to its many advantages, e.g., minimum operation and maintenance requirements/costs, and low excess sludge production. Anaerobic ammonium oxidation (anammox) is an anaerobic technique that converts ammonium into N2 where nitrite plays as the electron acceptor (Eq. (1)) (Abbas et al., 2015; Carvajal-Arroyo et al., 2014; Strous et al., 1998; Tao et al., 2012). Compared with the traditional nitrification/denitrification process, exogenous oxygen or organic compounds are dispensable in the anammox process, making it more cost efficient. NH4 + + 1.32NO2 − + 0.066HCO3 − + 0.13H+ → 1.02N2 + 0.256NO3 − + 0.066CH2 O0.5 N0.15 + 2.03H2 O

(1)

∗ Corresponding author at: College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036, China. E-mail address: [email protected] (R.-C. Jin). http://dx.doi.org/10.1016/j.ecoleng.2016.04.004 0925-8574/© 2016 Elsevier B.V. All rights reserved.

The anaerobic baffled reactor (ABR), which has been evolving since the early 1980s, is known as a highly efficient anaerobic reactor configuration that has many advantages compared with other reactors, e.g., longer retention times, less vulnerable to shock loadings, and the ability to separate the anaerobic metabolic phases (Barber and Stuckey, 1999; Shanmugam and Akunna, 2010). An ABR is a series of upflow anaerobic sludge blanket (UASB) reactors, and each compartment (C) is separated by a vertical baffle, which forces the wastewater flow up and down. The wastewater comes in close contact with the active biomass as it passes through the ABR Cs. It has been recently reported that ABRs are appropriate for the treatment of nitrogenous compounds contained wastewater (Jin et al., 2013a; Uyanik, 2003; Wang et al., 2004; Wu et al., 2013). However, owing to the compartmentalized structure, the initial Cs of the ABR may be overloaded with substrate, whereas the final Cs would be nutrient limited (Uyanik, 2003). To obtain a robust operation, several researchers have attempted to find new ways to optimize the stability and capacity of ABRs, such as combination utilization and operational optimization. Some researchers have attempted to combine the anaerobic and aerobic processes, such as by making an ABR-aerobic biofilm reactor (Bodik et al., 2003) and incorporating an anaerobic stage followed by an aerobic one in a modified ABR

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(Barber and Stuckey, 2000). Lu et al. (2011)investigated the effectiveness of the combined technique with an ABR-sequencing batch reactor (SBR) system for hypersaline wastewater treatment and obtained satisfactory results. Uyanik (2003) employed two feeding regimes and found that a split-fed ABR has a number of potential advantages over the normally fed ABR during both the start-up period and steady operation period. A flow switched anaerobic baffled reactor (FSABR) was proposed to eliminate the technical problems of substrate confusion in the Cs. Long-term substrate over-loading and nutrient-limited situations can be avoided by periodically switching the feeding direction. The objective of this study was to treat nitrogen-containing wastewater with an anammox FSABR with different feeding regimes and to test the superiority of a flow-switched feeding regime over the normal regime in terms of the reactor potential, robustness, and sludge characteristics. 2. Materials and methods 2.1. Seed sludge and anammox reactor An FSABR was used in this study and its working volume was 4 L (Fig. 1). The reactor was constructed with a height of 32 cm, width of 6.2 cm, bottom length of 36 cm and top length of 51.2 cm. The reactor was inoculated with mature anammox sludge from a sturdily operated anammox reactor in the laboratory. The suspended solids (SS) concentration of the sludge after inoculation was 10.4 gL−1 with a volatile suspended solids (VSS) concentration of 6.9 g L−1 . The reactor was observed for 354 days in the thermostatic room where the temperature was35 ± 1 ◦ C, and the research could be divided into two stages: a CABR from days 1 to 220 (Fig. 1A) and a FSABR from days 221 to 354, during which the feeding regime was alternated between the two feeding regimes every 10 days (Fig. 1A and B).

Extracellular polymeric substances (EPS) were extracted by ‘heating’ method, and the method described by Ma et al. (2012) was utilized to measure polysaccharide (PS) and extracellular protein (PN) concentrations. Heme c was measured by the method used by Berry and Trumpower (1987). Specific anammox activity (SAA) assays were measured in batch assays according to Yang and Jin (2013). 2.4. Statistical analysis Statistical comparison between variables was conducted using One-Way analysis of variance (ANOVA) and the Mann-Whitney Test by SPSS software (SPSS 13.0). A p-value of 0.05 or lower indicates that the difference between the variables under comparison is statistically significant. 2.5. Stover-Kincannon model The Stover-Kincannon model has been widely applied in immobilized systems in order to calculate the kinetic constants (Isik and Sponza, 2005; Jin and Zheng, 2009; Stover and Kincannon, 1982). In this study, this model was modified and used in the FSABR (Eq. (2)).

 dS -1 dt

=

V 1 KB V + = Q (S0 -S) Umax QS0 Umax

(2)

in which dS/dt is the substrate removal rate (kg m−3 d−1 ); Umax is the maximum substrate removal rate (kg m−3 d−1 ); Q is the inflow rate (L d−1 ); S0 and S are influent and effluent total nitrogen concentrations (kg m−3 ); V is the reactor volume (L) and KB is the saturation value constant (kg m−3 d−1 ). 3. Results

2.2. Synthetic wastewater

3.1. Anammox performance at different feeding regimes

The synthetic wastewater contained substrates, inorganic solution and trace elements. (NH4 )2 SO4 and NaNO2 were used as the source of ammonium and nitrite, respectively. Trace elements, including EDTA at 15 g L−1 , CoCl2 ·6H2 O at 0.24 g L−1 , CuSO4 ·5H2 O at 0.25 g L−1 , ZnSO4 ·7H2 O at 0.43 g L−1 , MnCl2 ·4H2 O at 0.99 g L−1 , NiCl2 ·2H2 O at 0.19 g L−1 , NaMoO4 ·2H2 O at 0.22 g L−1 , H3 BO4 at 0.014 g L−1 , NaSeO4 ·10H2 O at 0.21 g L−1 , and NaWO4 ·2H2 O at 0.050 g L−1 , were supplied at a dose of 1.25 mL per liter of wastewater (Yang and Jin, 2013). The inorganic solution consisted of NaH2 PO4 at 10 mg L−1 , CaCl2 at 5.65 mg L−1 , KHCO3 at 1 g L−1 and MgSO4 ·7H2 O at 58.6 mg L−1 .

3.1.1. Stage 1 (days 1–220) In stage 1, the reactor was operated as a CABR. The supplementation of NO2 − -N and NH4 + -N was in a ratio of 1:1 which was demonstrated appropriate for the anammox reactor according to our previous studies (Jin et al., 2013a,b,c; Ma et al., 2012). The hydraulic retention time (HRT) was initially set at 21.3 h, and the initial concentrations of NO2 − -N and NH4 + -N were 70 mg L−1 , and high nitrogen removal efficiency (NRE) was achieved. Thus, the HRT was shortened to 4.1 h within 10 days for a given substrate concentration. The NRE consistently remained above 99.0% during these days. As shown in Fig. 2, from days 8 to 75, the HRT was consistently maintained at 4.1 h, and the concentration of the substrate gradually increased. On day 37, the concentration was 308 mg L−1 , and the effluent NO2 − -N started to ascend gradually over 100 mg L−1 (day 41), which may lead to the risk of nitrite inhibition to the anammox system (Strous et al., 1999). Hence, the concentration was decreased until the anammox system recovered at 70 mg L−1 on day 73. During days 8–75, a maximum nitrogen removal rate (NRR) of 3.00 kgN m−3 d−1 was obtained. After the recovery of the anammox system, the influent substrate concentrations were fixed, and an exploration of the minimum HRT that could be achieved by the CABR was conducted. The HRT was gradually shortened until the HRT reached 3.3 h. The nitrogen conversion decreased due to the accumulation of nitrite. To obtain a maximum NRR in the rest time of stage 1, both the HRT and substrate concentrations were adjusted. However, no NRR greater than 3.00 kgN m−3 d−1 was achieved.

2.3. Analytical methods 25 mL effluent were sampled with a small beaker every 2 days, and the samples were stored in a 4 ◦ C refrigerator for the measurement. The pH was measured with a pH meter (Mettler Toledo Delta 320, Switzerland), SS and VSS was measured by weight method, and NO3 − -N, NO2 − -N, and NH4 + -N were measured by spectrophotometry. The pH, SS, VSS, NO2 − -N, NO3 − -N, and NH4 + -N were determined with the methods described in standard methods (APHA, 2005). The particle diameter was measured by the method described by Jin et al. (2013b). The settling velocity (VS ) was determined in a measuring cylinder (1 L), the height and internal diameter of the cylinder was 40.0 cm and 10.0 cm in order to minimize the wall effect. The sludge was slightly put into the middle of the water surface. When the sludge reach the line of 750 mL the timer starts and the time for the distance of 20 cm are recorded.

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Fig. 1. Configuration and feeding regimes of the FSABR. (A) fed from the left side; (B) fed from the right side.

3.1.2. Stage 2 (days 221–354) To optimize the performance of the CABR, the FSABR was employed in stage 2. In this stage, the influent flow direction was switched every 10 days. The performance of the FSABR is shown in Fig. 2C and D. At the beginning of this stage, the effluent NO2 − N concentration reached 60 mg L−1 ; the substrate concentration and HRT were changed to 140 mg L−1 and 9.0 h, respectively. Similar to stage 1, the nitrogen loading rate (NLR) was increased by increasing the substrate concentration or shortening the HRT. In summary, a minimum HRT of 2.7 h was obtained when the influent substrate concentration was fixed at 210 mg L−1 , and 364 mg L−1 was achieved when the HRT was 4.1 h. In addition, a maximum NRR of 3.49 kgN m−3 d−1 was obtained in stage 2.

Table 1 Indices of the FSABR during the two stages. Stage 1

−3

−1 a

NLR (kgN m d ) NRR (kgN m−3 d−1 )b NRE (%)c RS d RP e *

Stage 2

Average

Std.*

Average

Std.*

1.62 1.24 75.4 1.20 0.18

0.77 0.66 15.7 0.47 0.10

1.80 1.51 83.1 1.32 0.26

1.03 1.09 13.7 0.26 0.08

Standard deviation. a Nitrogen loading rate. b Nitrogen removal rate. c Nitrogen removal efficiency. d Consumption NO2 − -N to consumption NH4 + -N. e Production NO3 − -N to consumption NH4 + -N.

3.2. Stoichiometric ratio at different feeding regimes The stoichiometric ratio is a constant value that is used to calculate the relationship between products and reactants in a biochemical reaction (Strous et al., 1998). The stoichiometric ratios of the reactor are listed in Table 1. The production of NO3 − -N to the consumption of NH4 + -N (RP ) and the consumption of NO2 − -N to NH4 + -N (RS ) were 0.18 ± 0.10 and 1.20 ± 0.47 in the CABR, and the RS and RP values in the FSABR were 1.32 ± 0.26 and 0.26 ± 0.08, respectively. The theoretical stoichiometric ratios for RS and RP were 0.26 and 1.32, respectively (Strous et al., 1998). 3.3. Sludge characteristics at different feeding regimes Sludge characteristics have a direct relationship with the reactor performance; hence, some of the characteristics were summarized

in Table 2, and the details of the heme c are presented in the Supplementary materials. The biomass from the high-rate anammox reactor typically has a higher SAA value than that of the reactor with poor capacity (Yang and Jin, 2013). As shown in Table 2, the SAA gradually decreased from C1 to C4 on day 220, and after the operation of the flowswitched feeding regime, the corresponding SAA values increased by more than 5-fold. Good settling properties and proper size distribution of the granules are important for the retention of the biomass and the sturdy operation of the reactor (Jin et al., 2013c). In the present study, the VS are 62.5 ± 16.1, 62.9 ± 13.7, 55.0 ± 15.4 and 75.9 ± 15.1 m h−1 on day 220, and the values in the corresponding Cs are 85.4 ± 14.7, 70.4 ± 14.2, 51.6 ± 13.6 and 80.3 ± 21.7 m h−1 , respectively. The size

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Fig. 2. Performance of the FSABR throughout the study. (A) and (B) capacity of the reactor during stage 1; C and D, capacity of the reactor during stage 2.

Table 2 Comparison of the sludge characteristics of the biomass, including the SAA, EPS, heme c, and VS on days 220 and 354. Day 220

Day 354 *

*

C1 SAA (mgN h−1 g−1 VSS)a EPS (mg EPS g−1 VSS)b VS (m h−1 )e *

PNc PSd

1.26 189.6 100.9 62.5

*

C2 ± ± ± ±

0.17 11.4 8.8 16.1

0.66 142.3 93.3 62.9

C4*

C3 ± ± ± ±

0.15 13.1 6.5 13.7

0.76 133.9 80.0 55.0

± ± ± ±

0.10 10.3 7.2 15.4

0.45 143.6 74.5 75.9

C1* ± ± ± ±

0.08 9.9 4.6 15.1

7.95 287.2 176.8 85.4

C2* ± ± ± ±

0.77 18.9 16.8 14.7

6.73 308.4 179.0 70.4

C3* ± ± ± ±

0.45 15.6 20.1 14.2

4.15 265.7 94.7 51.6

C4* ± ± ± ±

0.52 22.3 8.7 13.6

4.11 260.3 133.1 80.3

± ± ± ±

0.34 23.2 12.1 21.7

Compartment. a Specific anammox activity. b Extracellular polymeric substances. c Protein. d Polysaccharide. e Settling velocity.

distribution of the granules is shown in Fig. 3. In stage 1, the particles were mainly small, especially in C3, where 86.5% of particles were smaller than 1 mm. However, in stage 2, the size distribution was more homogeneous.

4. Discussion 4.1. Differences in the performance between the CABR and FSABR The operational conditions during the two stages were identical; however, the capacity of the FSABR was better than the CABR (83.1 ± 13.7% vs. 75.4 ± 15.7% in NRE (p < 0.01, Table 3), 1.80 ± 1.03 kgN m−3 d−1 vs. 1.62 ± 0.77 kgN m−3 d−1 in

NLR and 1.51 ± 1.09 kgN m−3 d−1 vs. 1.24 ± 0.66 kgN m−3 d−1 in NRR, respectively). In stage 1,a minimum HRT of 3.3 h was obtained at 70 mg L−1 , and the average NRE for this duration was 67.7 ± 13.1%. A maximum substrate concentration of 308 mg L−1 was achieved when the HRT was 4.1 h, and the average NRE was 80.2 ± 4.5%. As a contrast, the minimum HRT of 2.7 h was obtained in stage 2 at 210 mg L−1 , and the average NRE was 89.1 ± 2.2%. Furthermore, the NRE of the FSABR was 88.3% when the substrate concentration and HRT were 336 mg L−1 and 4.1 h, respectively. The reactor performance was evidently improved after the feeding regime change. An ABR is a specially designed configuration with near plugflow characteristics (Krishna et al., 2008; Skiadas et al., 2000); thus, the substrate concentration in the reactor will gradually decrease

H. Chen et al. / Ecological Engineering 92 (2016) 229–235

233

Fig. 3. Size distribution in each compartment on days 220 and 354.

Table 3 Summary of the statistical analysis of the FSABR. Parameter

Significance

Model

NREa (%) SAAb EPS

0** 0.021* 0.043* 0.021* 0.386 0** 0**

One-way ANOVA Mann-Whitney Test Mann-Whitney Test

PSc (mg g−1 VSS) PNd (mg g−1 VSS)

Heme c (␮g g−1 VSS) VS e (m h−1 ) Diameter (mm) * ** a b c d e

Mann-Whitney Test Mann-Whitney Test Mann-Whitney Test

p < 0.05. p < 0.01. Nitrogen removal efficiency. Specific anammox activity. Polysaccharide. Proteins. Settling velocity.

from the inlet to the outlet. Therefore, the anammox biomass in the first C may suffer from long-term overloaded substrate feeding (Uyanik, 2003). Thus, there may be compound accumulation, such as the NO2 − -N, which is adverse for the biomass. Numerous studies have reported that nitrite concentration is a key factor to maintain the stability of the anammox reactors, and nitrite can lead to severe inhibition in some cases (Dapena-Mora et al., 2007; Jin et al., 2012; Kimura et al., 2010). When operated as a CABR, the anammox biomass in the former Cs has more opportunities to be influenced by long-term substrate overloading. In this study, changing the CABR to FSABR was successful in improving the reactor performance. Because the flow direction was switched every 10 days, the biomass was not exposed to substrate overloading for a long time, and therefore, the inhibition caused by NO2 − -N was eliminated in the FSABR. By contrast, when the FSABR was in good condition, the substrates were completely removed far before they reached the latter Cs. Hence, the biomass in the latter Cs would suffer from substrate starvation. The FSABR could avoid this situation and ensure the SAA of the biomass by protecting the Cs from long-term starvation. Moreover, in the FSABR, the biomass did not exhibit long-term exposure to substrate overloading or starvation. Similar results were obtained by other researchers by modifying the feeding mode or reactor configuration (Uyanik, 2003; Zwain et al., 2013). Zwain et al. (2013) found that a high COD removal efficiency

was obtained during the start-up operation of a modified ABR (up to 71% on day 30). Uyanik (2003) confirmed the advantages of the split feeding ABR over the normal feeding ABR by the reduction in the severity of conditions (toxicity) in the initial Cs of the reactor and provision of supplementary substrates to the biomass in the final C of the reactor. As previously stated, the theoretical stoichiometric ratios of anammox were 1.32 (RS ) and 0.26 (RP ) (Strous et al., 1998). The RS and RP values in stage 1 and 2 are listed in Table 1 (1.20 ± 0.47 vs. 1.32 ± 0.26 for RS and 0.18 ± 0.10 vs. 0.26 ± 0.08 for RP ). It is evident that the values in stage 1 deviated from the theoretical ratios; meanwhile, the NRE during stage 1 was only 75.4 ± 15.7%. Perhaps while operating as a CABR, the dominant organisms included both anammox bacteria and some other types of bacteria, such as ammonium oxidation bacteria (AOB); thus, the stoichiometric ratios in stage 1 deviated from the theoretical values. In the presence of AOB, a considerable part of NH4 + -N was converted to NO2 − -N, and it led to the insufficiency of NH4 + -N in the anammox reaction and less NO3 − -N was produced. Therefore, the RS and RP values decreased. When the feeding regime switched, the FSABR resulted in a good performance because the absence of long-term substrate overloading of the anammox biomass in the Cs. Hence, the anammox consortia became dominant again and the stoichiometric ratios and the NRE recovered. The stoichiometric ratios are the evidences for the stability of the anammox system and the evolution of the microbial community (Sun et al., 2012). The ratios will be influenced by many factors, such as the substrates, operating conditions and the reactor configurations. Therefore, the ratios will deviate under some stressing factors. Ma et al. (2012) conducted the trials under transient NaCl shocks. The results revealed that the ratios were easily shifted under the shocks. NO2 − -N is toxic to a wide variety of microorganisms, including anammox bacteria. In the center of enzymes, NO2 − -N is very reactive against biomolecules and it has a high affinity for the metals (Carvajal-Arroyo et al., 2014; Philips et al., 2002). Long-term exposure to NO2 − -N may lead to negative effects due to the accumulation of NO2 − -N in the sensitive area inside the cells (e.g., anammoxosome, riboplasm) (Carvajal-Arroyo et al., 2013) and eventually, change the stoichiometric ratios.

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Table 4 EPS contents in different studies. Sludge

Anammox granules Aanerobic granules Denitrifying granules Aerobic granules Anammox granules (stage 1) Anammox granules (stage 2) a b c d

EPSa (mg g−1 VSS)

References

PNb

PSc

PN/PS

164.4 ± 9.3 42.7 ± 37.8 N.G.d ≈40 150.5 ± 26.3 280.4 ± 22.0

71.8 ± 2.3 17.6 ± 6.8 N.G.e ≈16 87.2 ± 12.1 145.9 ± 40.2

2.29 2.50 2.20 2.50 1.73 1.92

[35] [41] [40] [42] This paper

Extracellular polymeric substances. Proteins. Polysaccharide. Not given.

4.2. Changes in sludge characteristics Generally, the sludge characteristics, including the SAA, EPS, VS , particle diameters, and heme c, are important parameters, which usually keep in accordance with the reactor capacity (Yang and Jin, 2013). Tang et al. (2011) reported that the high activity of the anammox biomass was an important factor to achieve the super high NRR in the UASB reactors. The fact that the SAA of the biomass increased considerably after the switch of feeding regime, which resulted in the improvement of NRR from 1.24 to 1.51 kgN m−3 d−1 (p < 0.05) as obtained in this study, is in accordance with the results achieved by Tang et al. (2011). Good settleability of the anammox granule ensures the process stability (Jin et al., 2012). The formation of granules with good settleability could ensure the retention of biomass in the reactor, and it was another factor contributing to the enhancement of the NRR and NRE. In this study, the average VS increased from 64.1 m h−1 in stage 1–71.9 m h−1 in stage 2 (p < 0.01), and the corresponding NRR increased from 1.24 to 1.51 kgN m−3 d−1 . In other words, the VS in this study had a positive relation with the NRR, which corresponded with the results achieved by Tang et al. (2011). Moreover, not only the VS but also the particle diameters positively related with the NRR. In stage 1, the CABR was dominated by small particles with an average diameter of 0.8 mm, and more than 70% particles were smaller than 1.0 mm. After the cultivation in the FSABR, the average diameter increased to 2.3 mm (p < 0.01), and the proportion of particles smaller than 1.0 mm decreased to 27.3%. Wang et al. (2012) reported the influence of the morphology and size of the granules on the microbial activity. Additionally, Vlaeminck et al. (2010) reported that the granules in the range of 0.5–1.6 mm, which is similar to the range achieved in this study, could remove nitrogen efficiently. The EPS is critical during sludge granulation in bioreactors (Liu and Tay, 2002; Liu et al., 2009). Along with the aggregation of particles, the EPS contents in the granule are increased. Both PN and PS increased in the anammox biomass when the FSABR was switched from the CABR with a p-value below 0.05 (Table 2 and 3). The proteins to polysaccharides ratio (PN/PS) can be used to describe the granular settleability (Franco et al., 2006; Tang et al., 2011; Wu et al., 2009). Franco et al. (2006) found that anaerobic granules possessing higher PN/PS ratio had a lower settleability, and the PN/PS ratios in this study (1.73 and 1.92 in stages 1 and 2, respectively) were lower than those in other studies (Table 4), suggesting a greater settleability. Wu et al. (2009) noted that the fluid viscosity in the reactor will be raised due to the over-production of EPS; thus, the shear force increases in the reactor based on Newton’s law. This increased shear force can make the sludge easier to be disrupt, and sludge washout becomes inevitable. As described by Chen et al. (2012), a number of factors can influence the morphology of the microbial granules, including the

Fig. 4. Modified Stover-Kincannon model plots.

heme c. The denitrifying granules and nitrifying granules are typically cream-colored, whereas the presence of sulfides makes the methanogenic granules black (Franco et al., 2006; Liu et al., 2009). The mature anammox granules are uniquely camine, and the heme c content is presumed to play a key role in the camine color of the granules. In this study, there is only a slight change in the composition of heme c (p = 0.386). Therefore, the colors of the granules in the two stages were similar.

4.3. Stover-Kincannon model Fig. 4 presents a plot of the reciprocal of the NRR, V/[Q(S0 -S)], against the reciprocal of the NLR, V/(QS0 ). According to Eq. (2), KB and Umax were calculated as 2.11 and 4.28 kg m−3 d−1 , respectively, in stage 1, whereas the corresponding values were 25.96 and 46.38 kg m−3 d−1 , respectively, in stage 2 with high correlations (0.9238 and 0.9397, respectively). Table 1 presents the average NRRs during the two stages. There is a large difference between the Umax and the results obtained in this study. Moreover, the stimulations indicated that the Umax in stage 2 increased by 10-fold compared with that in stage 1. The stimulation results were in line with the enhancement of characteristics discussed above. In summary, the FSABR had a more promising capacity than the CABR, and the feeding regime could be an effective way to enhance the reactor performance.

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In this study, the FSABR obtained a stable NRE of 83.1% and a maximum NRR of 3.49 kgN m−3 d−1 , and the performance was superior to that of the CABR; meanwhile, the sludge characteristics, including the SAA, EPS, VS , and particle diameter, were also enhanced from the CABR to the FSABR. The Stover-Kincannon model was applied for the FSABR, resulting in a high correlation coefficient in each stage (0.9238 and 0.9397 for stages 1 and 2, respectively). Similar results were achieved in other studies where the Stover-Kincannon model was used (Isik and Sponza, 2005; Jin and Zheng, 2009; Stover and Kincannon, 1982). Therefore, the performance of the FSABR can be predicted with the model. 5. Conclusion The FSABR obtained a stable NRE of 83.1% and a maximum NRR of 3.49 kgN m−3 d−1 under a flow-switched feeding regime, and the performance was superior to that under a normal feeding regime. The performance of the FSABR was improved when the normal feeding regime was changed to the flow-switched feeding regime; meanwhile, the sludge characteristics, including the SAA, EPS, VS , and particle diameter, were also enhanced. However, the heme c content changed only slightly. The results obtained were statistically analyzed in this study, and the changes in the reactor performance and sludge characteristics, except for the heme c, were found to be statistically significant. The Stover-Kincannon model was applied for the FSABR, resulting in a high correlation coefficient in each stage (0.9238 and 0.9397 for stages 1 and 2, respectively). Therefore, the performance of the FSABR can be predicted with the model. Acknowledgments The authors wish to thank the Natural Science Foundation of China (Nos. 51278162 and 51578204) and the Science and Technology Development Program of Hangzhou (No. 20120433B20) for their partial support of this study. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.ecoleng.2016.04. 004. References American Public Health Association (APHA), 2005. American Water Works Association (AWWA), Water Environment Federation (AEF), 2005 Standard Methods for the Examination of Water and Wastewater, 21st ed., Washington, DC, USA. Abbas, G., Wang, L., Li, W., Zhang, M., Zheng, P., 2015. Kinetics of nitrogen removal in pilot-scale internal-loop airlift bio-particle reactor for simultaneous partial nitrification and anaerobic ammonia oxidation. Ecol. Eng. 74, 356–363. Barber, W.P., Stuckey, D.C., 1999. The use of the anaerobic baffled reactor (ABR) for wastewater treatment: a review. Water Res. 33, 1559–1578. Barber, W.P., Stuckey, D.C., 2000. Nitrogen removal in a modifiedanaerobic baffled reactor (ABR): 2 nitrification. Water Res. 34, 2423–2432. Berry, E.A., Trumpower, B.L., 1987. Simultaneous determination of hemes a, b, and c from pyridine hemochrome spectra. Anal. Biochem. 161, 1–15. Bodik, I., Kratochvil, K., Gaspirkova, E., Hutnan, M., 2003. Nitrogenremoval in an anaerobic baffled filter reactor with aerobic post-treatment. Bioresour. Technol. 86, 79–82. ˜ A., Sierra-Alvarez, R., Field, Carvajal-Arroyo, J.M., Puyol, D., Li, G., Lucero-Acuna, J.A., 2013. Pre-exposure to nitrite in the absence of ammonium strongly inhibits anammox. Water Res. 48, 52–60. Carvajal-Arroyo, J.M., Puyol, D., Li, G.B., Sierra-Álvarez, R., Field, J.A., 2014. The role of pH on the resistance of resting- and active anammox bacteria to NO2 − inhibition. Biotechnol. Bioeng. 111, 1949–1956. Chen, T.T., Zheng, P., Shen, L.D., 2012. Growth and metabolism characteristics ofanaerobic ammonium-oxidizing bacteria aggregates. Appl. Microbiol. Biotechnol. 97, 5575–5583. Dapena-Mora, A., Fernandez, I., Campos, J.L., Mosquera-Corral, A., Méndez, R., Jetten, M.S.M., 2007. Evaluation of activity and inhibition effects on Anammox

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