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Differentiating two partial nitrification mechanisms: Inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity
Accepted Manuscript Title: Differentiating two partial nitrification mechanisms: Inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity Author: Jun Wu Yue Zhang Gangyan PII: DOI: Reference:
S2213-3437(16)30248-2 http://dx.doi.org/doi:10.1016/j.jece.2016.06.037 JECE 1170
To appear in: Received date: Revised date: Accepted date:
18-3-2016 7-6-2016 30-6-2016
Please cite this article as: Jun Wu, Yue Zhang, Gangyan, Differentiating two partial nitrification mechanisms: Inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity, Journal of Environmental Chemical Engineering http://dx.doi.org/10.1016/j.jece.2016.06.037 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Differentiating two partial nitrification mechanisms: inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity
Jun Wu*, Yue Zhang, Gangyan
School of Environmental Engineering and science, Yangzhou University, 196 West Huayang Road, Yangzhou, Jiangsu, 225127, China. Tel: +86-514-87971389, Fax: +86-514-87978626
*Correspondence author Email address: [email protected]
1
Highlights
Partial nitrification was closely associated to the specific biomass NH4+ load;
A low SRT was needed to maintain high specific biomass NH4+ load;
FA inhibition of NOB was insignificant for partial nitrification;
Increased AOB activity was the main reason for partial nitrification ;
Abstract Partial nitrification is usually more easily achieved for high NH4+ concentration wastewater. In this study, a two-compartment aerobic nitrification reactor was used to raise the NH4+ concentration in the first aerobic compartment and to enhance partial nitrification for low NH4+ concentration wastewater. In addition to the regular sludge wastage to maintain sludge retention time (SRT), extra sludge wastage was used to increase the specific biomass NH4+ load. The experimental results indicated that partial nitrification was closely associated to the specific biomass NH4+ load. The modelling results indicated that a low SRT was needed to maintain the low biomass concentration and high specific biomass NH4+ load. The increased AOB (ammonium oxidizing bacteria) activity, instead of the free ammonia (FA) inhibition of NOB (nitrite oxidizing bacteria) was identified to be the main mechanism for partial nitrification in low NH4+ concentration wastewater treatment with high specific biomass NH4+ load.
Keywords: nitritation; sludge retention time; ammonium load; two-compartment nitrification
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1. Introduction In the partial nitrification (nitritation), the ammonium (NH4+) oxidation was controlled at nitrite (NO2-) step, without being further oxidized into nitrate (NO3-) [1]. The partial nitrification offers several distinctive advantages including oxygen saving in the nitrification process, reduced carbon source requirement in the denitrification process and less sludge production [2]. Partial nitrification is also a requirement for the application of the recently developed autotrophic nitrogen removal by anaerobic ammonium oxidization (ANAMMOX) [3]. Applying oxygen limitation is one of the most widely used techniques for partial nitrification [4]. The rationale behind this operation is that NOB (nitrite oxidizing bacteria) has lower DO (dissolved oxygen) affinity than AOB (ammonium oxidizing bacteria) [5, 6]. Therefore, NOB will be washed out under DO limitation. However, recent studies have indicated the certain species of NOB could have higher DO affinity than AOB [7, 8], therefore, applying low DO will cause more inhibition in AOB activity than in NOB activity. Furthermore, the effect of low DO on partial nitrification is also complicated by other NOB inhibiting factors. Free ammonia (FA) inhibition of NOB was considered to another measure or achieving partial nitrification [9]. In studies that relied on FA inhibition on NOB for partial nitrification operation, DO limitation was also applied for a combined inhibition on NOB by FA and low DO [10, 11]. It is usually difficult to differentiate the exact contribution of NOB inhibition by low DO or FA. To elucidate the exact contribution of partial nitrification by FA, the DO was supplied in non-limiting condition in this study. Partial nitrification is usually assumed to be more easily achieved for high NH4+ load condition [12]. The FA inhibition on NOB was thought to be the main reason; however, besides reducing NOB activity by FA inhibition, the contribution of high NH4+ concentration on NO2- accumulation can also be explained by the increased AOB activity due to the high Monod term value for NH4+ [13]. No studies have been carried out to elucidate the quantitative contribution of high NH4+ concentration on NO2- accumulation. Partial nitrification for low NH4+ concentration wastewater (less than ca. 100 mg N/L) is an important step to implement the recently developed ANAMMOX technique in the municipal wastewater treatment [14]. The bulk liquid NH4+ concentration in the conventional CSTR (continuously stirred tank reactor) for municipal wastewater treatment is usually too low to take advantage of the high NH4+concentration for partial nitrification. To overcome the problem, the aerobic reactor was split into two compartments. It was expected that the high NH4+ in the first compartment could enhance partial nitrification by decreasing the NOB activity (by FA inhibition), or increasing the AOB activity (by increasing the value of Monod term for NH4+). 3
Mathematical model was included to elucidate the exact mechanisms of partial nitrification in the two-compartment reactor. 2. Materials and methods 2.1. Experimental bioreactor The schematic layout of the experimental bioreactor was shown in Fig.1. It includes one anoxic compartment and two aerobic compartments (#1, #2). Each compartment has an effective volume of 2 L. The effective volume of the settlement tank is 1.5 L. The influent flow rate (Q) was maintained at 0.5 L/h, resulting hydraulic retention time (HRT) of 12 h. The recirculation flow ratio (R1 and R2) from #1 and #2 aerobic compartment to the anoxic compartment was varied according to experimental plan shown in Table-1. The sludge retention time (SRT) was adjusted by wasting 400 ml of mixed liquor from the #2 aerobic compartment. As the solids concentration in the #2 aerobic compartment was roughly twice as high as the solids concentration in the other two compartments, the effective SRT was calculated to be 10 d. Synthetic wastewater was prepared according the recipe from Wu & He [15]. The chemical oxygen demand (COD) and N-NH4+ ratio was maintained at above 6 to allow complete denitrification in the anoxic tank. The seeding sludge was taken from the aeration tank of the local wastewater treatment plant. The bioreactor was placed in an air-conditioned room with temperature controlled at 25.0±0.8 ºC. The DO concentration in both of the aerobic compartments was provided at above 6 mg O2/L to create an oxygen non-limiting condition. 2.2. Experimental plans Before taking samples for measurement, the bioreactor was operated for 20 days of assimilation period. Then the bioreactor was operated according to the experiment plan shown in Table 1. During the experiment period, the NH4+, NO2- and NO3concentration in the influent, anoxic compartment and two aerobic compartments were measured regularly. The pH and NH4+ concentration in the influent were varied to examine the effect of FA on NOB inhibition. The changes in pH, influent NH4+ concentration and recirculation ratio were also necessary to provide enough data excitation for the mathematical model calibration [16]. At the end of period A, in addition to the regular sludge wastage to maintain the SRT, an extra portion of biomass solids was wasted to further decrease the biomass concentration and increase the specific biomass NH4+ loading rate. This allowed the high NH4+ loading conditions to be investigated for low NH4+ concentration wastewater. 4
2.3. Batch experiment for the measurement of the maximum specific growth rate for AOB and NOB The oxygen uptake rate (OUR) was measured to calculate the maximum specific growth rate for AOB and NOB (μAOB and μNOB , d-1). The OUR was measured using the respirometer shown by Wu et al [17]. The OUR measured at time t under ammonium and DO non-limiting condition (OURNH(t)) can be expressed by the following equation [18]: OUR
NH
(t ) e
t(
AOB
b AOB )
OUR
0
(1)
NH
OURNH(t) is expressed by the exponential function of
OUR
0 NH
(the initial OUR after
ammonium addition). By curve fitting the measured OUR to the simulated OUR and assuming the decay rate bAOB to be 5% of μAOB , μAOB can be estimated. Similarly, the μNOB can be estimated by the following equation: OUR
NO 2
(t ) e
t(
NOB
b NOB )
OUR
0
(2)
NO 2
Where, OURNO2(t) is the OUR at time t after nitrite addition;
OUR
0 NO 2
is the initial
OUR after nitrite addition. 2.4. Sample analysis method The pH and DO were measured by HACH pH electrode and LDO oxygen sensor respectively. VSS, COD, NH4+, NO2- and NO3- concentration were measured according to the standard method [19]. 2.5. Mathematical modelling 2.5.1. Model kinetics and parameters The ASM (activated sludge model) type of model was included to simulate the reactor shown in Fig. 1 [16]. The model had 4 particulate components including AOB (XAOB), NOB(XNOB), heterotrophic bacteria (XH) and inert material (XI); and 6 soluble components including DO (SO), NH4+ (SNH), NO2- (SNO2) and NO3- (SNO3) and soluble COD (Ss). The complete model stoichiometric matrix and kinetic rate expressions were shown in the supporting information Table S1. The maximum specific growth rate for AOB and NOB were determined by respirometer shown in section 2.3. The FA inhibition constant for NOB (KIA) was determined from model calibration. Other parameter values was taken from the literature and shown in the Table S2. The growth rates AOB or NOB were expressed by the following equation: 5
dX
AOB
dt dX
NOB
dt
S AOB
K
S
S , NH
S NOB
K
SO
NH NH
K
S , NO 2
X
NO 2
K
O , NOB
(3)
AOB
SO
NO 2
S
SO
O , AOB
K SO K
IA
IA
FA
X
(4) NOB
Where, KS,NH and KS,NO2 were substrate half saturation constant for AOB and NOB; KO,AOB and KO,NOB were substrate half saturation constant for AOB and NOB; KIA was the FA inhibition constant. 2.5.2. Model calibration The NH4+, NO2- and NO3- concentration data measured during the experiment was used for model calibration. The KIA value was determined by ensuring the best curve fitting between the model prediction and experimental data. 2.4.3. Modelling strategy For the model based analysis, the influent was fixed at 50 mg N/L and pH was fixed at 7.5 to represent the municipal wastewater treatment. The simulation time was 300 days to reach steady state. The model was simulated under different combination of SRT and DO to investigate their effect on partial nitrification. To quantify the effect of FA on nitration, a new model that didn’t include the FA inhibition on NOB was run and compared to the original simulation results. In the new model, FA inhibition term for the NOB was removed. The NOB growth rate was expressed by the following equation: dX
NOB
dt
S NOB
K
SO
NO 2
S , NO 2
S
NO 2
K
O , NOB
SO
X
(5) NOB
Other kinetic rate expressions remained the same as shown in the supporting information Table S1. The AOB and NOB composition were compared for simulation results with different combination of SRT and DO. The amount of AOB was considered to be sufficient if more than 90% of NH4+ oxidation rate was achieved. The NOB washout was defined by NOB concentration of less than 0.1 mg COD/L. 3. Results and discussion 3.1. Reactor performance Fig. 2(a-c) showed the NH4+, NO2- and NO3- concentration monitored in the influent, anoxic and two aerobic compartments over the experiment periods specified in Table-1. Fig. 2(d) showed that the NAR (nitrite accumulation rate) in the two aerobic compartments, defined by the following equation:
6
NO
NAR NO
2
2
NO
(6)
100 %
3
The NO2- and NO3- concentration were near zero in the anoxic tank during the experiment periods (Fig.2(a)); therefore, a complete denitrification was achieved in the anoxic tank. During period A (day 0-23), the NH4+ was completely removed in both aerobic compartments (Fig.2(b-c)); the NO2- concentration gradually increased, whereas the NO3- concentration slightly decreased in the #1 aerobic compartment (Fig.2(b)); the NO2- was completely oxidized into NO3- in the #2 aerobic compartment (Fig.2(c)). Fig. 2(d) showed that the nitrite accmulation was low in the #1 aerobic compartment and remained at near zero in the #2 aerobic compartment during period A. In period B, the increase in influent NH4+ concentration and extra sludge solid loss of nearly 65% resulted in the high NH4+ accumulation up to 30 mg N/L in the #2 aerobic compartment (Fig. 2(b)). The measured VSS concentration in the bioreactor was shown in the Fig. S1. NO3- production was very low and nearly 100% nitrite accumulation was achieved in the #1 aerobic compartment (Fig. 2(b, d)). Despite the massive sludge loss, NH4+ accumulation in the #2 aerobic compartment was not significant (Fig. 2(c)). NO2- was initially accumulated and then decreased in the #2 aerobic compartment, with NAR increasing from ca. 60% to 100% initially and then deceasing to ca. 50% (Fig. 2(c, d)). In period C, the recirculation from the #1 aerobic reactor (R1) increased to 2.5 and the pH decreased to 8.2. The solid loss was recovered in the period C as demonstrated by the VSS measurement shown in Fig. S1. A low NO3- concentration was noticeable in the #1 aerobic compartment (Fig. 2(b)). The NO2- concentration decreased significantly in the #2 aerobic compartment (Fig. 2(c)). The NAR remained above 70% in the #1 aerobic compartment and deceased to near zero in the #2 aerobic compartment (Fig. 2(d)). Fig. 3 shows the specific biomass NH4+ loading rate and actual NH4+ removal (kg N/(kg VSS · d)) for both aerobic compartments. The specific biomass NH4+ loading rate was defined by the NH4+ loading per kg VSS per day (kg N/(kg VSS · d)). The sludge maximum NH4+ removal capacity (kg N/(kg VSS · d)) was derived from measurement of NH4+ removal at NH4+ and DO non-limiting condition in a batch reactor. During period A and C, nearly all the NH4+ load was removed in the #1 aerobic compartment. The NH4+ load entered the #2 aerobic compartment after the extra solids wastage. The maximum NH4+ removal capacity indicated that, except for the extra solid loss period, the system was operated at lower NH4+ loading rate than the NH4+ removal capacity. This explained why the NH4+ effluent concentration was not affected by the massive solid loss. 7
The high NAR (nitrite accumulation rate) was found to be closely associated with the NH4+ load. The high NAR in the first aerobic compartment could be explained by the higher NH4+ load compared to the second aerobic compartment. The sudden increasing NAR in the first aerobic compartment, following the solids loss, also pointed to the conclusion that increasing NH4+ load led to NO2- accumulation. The high NAR could not be sustained due to the recovering of biomass concentration and decreasing in the specific biomass NH4+ load. The maximum NH4+ removal capacity of ca. 1.2 kg N/(kg VSS · d) in this study was lower than the 2.87 kg N/(kg VSS · d) reported by Chen et al. [20], suggesting there was room for increasing the NH4+ load for NO2- accumulation. Because oxygen was provided in non-limiting condition, the results in Fig. 2 and Fig. 3 can suggest that NH4+ load was critical for NO2- accumulation. The result was consistent with previous reports in which NH4+ was provided in excess to promote stable partial nitrification [13]. However, these studies were carried out at extremely high NH4+ concentration (above ca. 1000 mg N/L). The result in this study indicated the partial nitrification could be achieved for low NH4+ concentration (less than ca. 100 mg N/L) by increasing the specific biomass NH4+ load via sludge wastage. The aggressive sludge wasting technique to achieve partial nitrification used by Regmi et al. [8] also supported the conclusion in this study. The NOB inhibition by FA under high NH4+ load was usually assumed to be the main mechanism for achieving partial nitrification [10]. On the other hand, AOB activity could be increased under high NH4+ load due to the increasing in Monod term for NH4+. The high AOB activity could also contribute to the NO2- accumulation. Although, the experiment result in this study showed that the high NH4+ load could promote NO2- accumulation. It is not sure whether it is due to the increased AOB activity or the decreased NOB activity. The modelling study was included to elucidate the mechanism. 3.2. Modelling results The maximum specific growth rates for AOB and NOB (μAOB, μNOB, d-1) were calibrated for the model shown in Table S1.The OUR measured under substrate non-limiting and high food to mass (F/M) ration condition used for measurement of μAOB and μNOB (d-1) was shown in Fig. 4 . Fig. 4(a) showed the OUR measured used for maximum specific AOB growth rate (μAOB, d-1) measurement under DO and NH4+ non-limiting condition. By curve fitting the OUR data to the equation 1, the μAOB value could be estimated at 0.98 d-1. Fig. 4(b) showed the OUR measured used for maximum specific NOB growth rate (μNOB, d-1) measurement under DO and NO2non-limiting condition. By curve fitting the OUR data to the equation 2, the μNOB value could be estimated at 0.62 d-1. The μAOB and μNOB value were higher than the normally used values [6], possibly due to the high NH4+ and NO2- load in the first 8
aerobic compartment. The high substrate availability will select fast growing r-strategist species [21]. 3.2.1. Description of the experimental results Fig. 5 showed the simulated and measured NH4+, NO2- and NO3- concentration in the first and second aerobic compartment. The initial AOB and NOB concentration and FA inhibition constant for NOB (KIA) were adjusted by trial and error method to produce the best fit between simulated and measured results. At day 24, the solids concentrations were reduced by 65% to represent the extra solid loss shown in Fig. S1. By choosing the KIA value at 0.2 (mg N/L), the simulation data was in agreement with the measured data. This confirmed that the validity of the model in represent the experimental results. The KIA value of 0.2 (mg N/L) was lower than reported by Jubany et al.[10]. The FA concentration between 0.1-1.0 mg N /L was suggested to be inhibitive to NOB [9]. Therefore, the KIA value was lower compared to the literatures. According to the equation 4, a low KIA value means a high FA inhibition of NOB. 3.2.2. Mechanisms of NO2- accumulation To quantify the effect of FA inhibition on NOB washout, simulations were carried out for model with and without FA inhibition term for NOB (equation 4 and 5). Fig. 6(a) showed the NOB washout or proliferation under different SRT and DO concentration. In general, low DO and SRT led to the NOB washout. The area below the curve marked with triangles represented NOB washout considering the FA inhibition of NOB. While the area below the curve marked with circles represented NOB washout without considering the FA inhibition of NOB. The area above the two curves represented the NOB proliferation. Compared to model without FA inhibition on NOB, the model with FA inhibition on NOB only predicts slight increasing in area that could be used for NOB washout (marked by grey shade in Fig. 6(a)). This suggested that the contribution of FA inhibition in NOB washout was insignificant for the bioreactor used in this study. It should be noted the KIA value of 0.2 (mg N/L) was lower than the values reported [10]. A low KIA value means strong inhibition. Even though, the contribution by FA inhibition for NOB washout was still insignificant. For KIA value of 0.95 (mg N/L) suggested by Jubany et al. [10], the simulation predicted a much smaller area of NOB washout contributed by FA inhibition (Fig. S2). Since the simulation result suggested a very small the contribution of FA inhibition on NOB washout, NO2- accumulation observed in experiment with high NH4+ load condition could be attributed to the increased AOB activity.
9
Fig. 6(b) showed that the SRT and DO combination (area marked with light blue) that could be used for partial nitrification operation predicted by model without FA inhibition on NOB. The area above the curve marked with squares represented condition where there was abundant AOB, defined by NH4+ oxidation rate of above 90%. The area below the curve marked with red circles showed NOB washout. The common area for the AOB abundant and NOB washout represented the SRT and DO combination required for partial nitrification (marked with light blue shade). For example, the SRT of 4 days and DO concentration of 1 mg O2/L at green dot represented one partial nitrification operation point. The low SRT predicted by the model corresponded to the massive solids loss in this study and aggressive sludge wastage used by Regmi et al. [8]. The low biomass concentration under low SRT led to high specific biomass NH4+ load, therefore, enhanced the AOB activity and resulted in NO2- accumulation. The model prediction also explained why the high NO2accumulation after extra solids loss was not sustainable under SRT of 10 days, since the solids concentration would recover and reduce the specific biomass NH4+ load. The 5-pointed red star represented full nitrification operation point and the blue square represented insufficient NH4+ oxidation. Partial nitrification achieved under high NH4+ load condition has been reported for high NH4+ concentration wastewater (ca. 500-1000 mg N/L) [10, 20, 22]. The partial nitrification achieved during massive solid loss in this study indicated that the specific biomass NH4+ load rate could be increased to achieve partial nitrification for low NH4+ concentration condition (less than ca. 100 mg N/L). However, due to the long SRT of 10 days used in the experiment, the high specific biomass NH4+ load rate could not be sustained after the recovery of biomass concentration. The simulation results indicated that a low SRT can be used to maintain the high specific biomass NH4+ load rate.
10
The increased AOB activity under high FA concentration has been reported for high NH4+ concentration wastewater (ca. 500 mg N/L)[20]. The experimental and modelling in this study confirmed that the FA inhibition was not significant in NOB washout; instead, it was the increased AOB activity under high NH4+ load condition that led to NOB washout in low NH4+ concentration wastewater. The findings provided a theoretical basis for using low SRT to achieve high specific biomass NH4+ load rate and partial nitrification in low NH4+ concentration wastewater. 4. Conclusions
Partial nitrification was closely associated to the specific biomass NH4+ load.
FA inhibition of NOB was insignificant in the two-compartment reactor for partial nitrification.
The increased AOB activity was identified to be the main mechanism of partial nitrification for low NH4+ concentration wastewater treatment with high specific biomass NH4+ load.
Acknowledgement The financial support from Natural Science Foundation of China (Grant number: 51478410) is highly appreciated. The authors also thank the support from the Yangzhou University high-end talent program.
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Reference [1] O. Turk, D.S. Mavinic, Preliminary assessment of a shortcut in nitrogen removal from wastewater, Canadian Journal of Civil Engineering, 13 (1986) 600-605. [2] S. Aslan, L. Miller, M. Dahab, Ammonium oxidation via nitrite accumulation under limited oxygen concentration in sequencing batch reactors, Bioresour. Technol., 100 (2009) 659-664. [3] M.S.M. Jetten, S.J. Horn, M.C.M. van Loosdrecht, Towards a more sustainable municipal wastewater treatment system, Water Sci. Technol., 35 (1997) 171-180. [4] Y. Ma, Y. Peng, S. Wang, Z. Yuan, X. Wang, Achieving nitrogen removal via nitrite in a pilot-scale continuous pre-denitrification plant, Water Res., 43 (2009) 563-572. [5] A. Guisasola, I. Jubany, J.A. Baeza, J. Carrera, J. Lafuente, Respirometric estimation of the oxygen affinity constants for biological ammonium and nitrite oxidation, J. Chem. Technol. Biotechnol., 80 (2005) 388-396. [6] U. Wiesmann, Biological nitrogen removal from wastewater, Adv. Biochem. Eng. Biotechnol., 51 (1994) 113-154. [7] R. Manser, W. Gujer, H. Siegrist, Consequences of mass transfer effects on the kinetics of nitrifiers, Water Res., 39 (2005) 4633-4642. [8] P. Regmi, M.W. Miller, B. Holgate, R. Bunce, H. Park, K. Chandran, B. Wett, S. Murthy, C.B. Bott, Control of aeration, aerobic SRT and COD input for mainstream nitritation/denitritation, 57 (2014) 162-171. [9] A.C. Anthonisen, R.C. Loehr, T.B. Prakasam, E.G. Srinath, Inhibition of nitrification by ammonia and nitrous acid, J. Water Pollut. Control Fed., 48 (1976) 835-852. [10] I. Jubany, J. Lafuente, J.A. Baeza, J. Carrera, Total and stable washout of nitrite oxidizing bacteria from a nitrifying continuous activated sludge system using automatic control based on Oxygen Uptake Rate measurements, Water Res., 43 (2009) 2761-2772. [11] G. Ruiz, D. Jeison, R. Chamy, Nitrification with high nitrite accumulation for the treatment of wastewater with high ammonia concentration, Water Res., 37 (2003) 1371-1377. [12] D. Wei, X. Xue, L. Yan, M. Sun, G. Zhang, L. Shi, B. Du, Effect of influent ammonium concentration on the shift of full nitritation to partial nitrification in a sequencing batch reactor at ambient temperature, Chem. Eng. J., 235 (2014) 19-26.
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[13] E. Isanta, C. Reino, J. Carrera, J. Perez, Stable partial nitritation for low-strength wastewater at low temperature in an aerobic granular reactor, Water Res., 80 (2015) 149-158. [14] T. Lotti, R. Kleerebezem, Z. Hu, B. Kartal, M.K. de Kreuk, C. van Erp Taalman Kip, J. Kruit, T.L.G. Hendrickx, M.C.M. van Loosdrecht, Pilot-scale evaluation of anammox-based mainstream nitrogen removal from municipal wastewater, Environ. Technol., 36 (2015) 1167-1177. [15] J. Wu, C. He, Effect of cyclic aeration on fouling in submerged membrane bioreactor for wastewater treatment, Water Res., 46 (2012) 3507-3515. [16] M. Henze, W. Gujer, T. Mino, M. van Loosdrecht, Activated Sludge Models: ASM1, ASM2, ASM2d and ASM3 IWA Publishing, London, 2000. [17] J. Wu, C. He, The effect of settlement on wastewater carbon source availability based on respirometric and granulometric analysis, Chem. Eng. J., 189-190 (2012) 250-255. [18] A. Ramdani, P. Dold, S. Deleris, D. Lamarre, A. Gadbois, Y. Comeau, Biodegradation of the endogenous residue of activated sludge, Water Res., 44 (2012) 2179-2188. [19] AWWA, Standard Methods for the Examination of Water and Wastewater, 21st ed., American Water Works Association, 2005. [20] J. Chen, P. Zheng, Y. Yu, Q. Mahmood, C. Tang, Enrichment of high activity nitrifers to enhance partial nitrification process, Bioresour. Technol., 101 (2010) 7293-7298. [21] A. Terada, S. Sugawara, T. Yamamoto, S. Zhou, K. Koba, M. Hosomi, Physiological characteristics of predominant ammonia-oxidizing bacteria enriched from bioreactors with different influent supply regimes, Biochem. Eng. J., 79 (2013) 153-161. [22] L.-Y. Chai, M. Ali, X.-B. Min, Y.-X. Song, C.-J. Tang, H.-Y. Wang, C. Yu, Z.-H. Yang, Partial nitrification in an air-lift reactor with long-term feeding of increasing ammonium concentrations, Bioresour. Technol., 185 (2015) 134-142.
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Figure captions:
Fig.1 – Schematic layout of the experimental bioreactor Fig. 2 – (a) Influent NH4+ concentration (NH4+ #0) and NH4+, NO2- , NO3- in the anoxic tank (#a); (b) NH4+, NO2- , NO3- concentration in the first aerobic compartment (#1); (c) NH4+, NO2- , NO3- concentration in the second aerobic compartment (#2); (d) Nitrite accumulation rate (NAR) in the first and second aerobic compartment Fig. 3 –NH4+ loading rate, removal and maximum NH4+ removal capacity in the (a) first and (b) second aerobic compartment Fig. 4 – OUR measurements used for the determination maximum specific growth rate of (a) AOB and (b) NOB Fig. 5 – Measured (M) and simulated (S) NH4+, NO2- , NO3- concentration in (a) first and (b) second aerobic compartment Fig. 6 – (a) NOB washout or proliferation under different SRT and DO concentration for model with or without FA inhibition on NOB; (b) SRT and DO combination (area marked with light blue) that can be used for partial nitrification operation predicted by model without FA inhibition on NOB.
14
Figures:
Fig.1 – Schematic layout of the experimental bioreactor
15
A
-
C
40
D
0
NH+4 #1
Solids loss
NO-2 #1
30
D
NO-3 #1
20
A
B C
10
+
20 0
10
20
30 40 Time (day)
50
60
0
0
10
20
30 40 Time (day)
NH+4 #2
30
NO-2 #2
B
D
C
NO-3 #2
20 10
0
10
-
A
+
0
20
50
60
(d)
NO2 accmulation rate (%)
(c) 40
-
-
N-NH4, N-NO2, N-NO3 (mgN/L)
+
-
B
(b) 40
-
80
N-NH4, N-NO2, N-NO3 (mg N/L)
100
60
(a)
NH+4 #a NO-2 #a NO-3 #a
-
N-NH4, N-NO2, N-NO3 (mgN/L)
NH+4 #0
120
30 40 Time (day)
50
100
60
C A
40
D
B
20 0
60
Aerobic #2 Aerobic #1
80
0
10
20
30 40 Time (day)
50
60
Fig. 2 – (a) Influent NH4+ concentration (NH4+ #0) and NH4+, NO2- , NO3- in the anoxic tank (#a); (b) NH4+, NO2- , NO3- concentration in the first aerobic compartment (#1); (c) NH4+, NO2- , NO3- concentration in the second aerobic compartment (#2); (d) Nitrite accumulation rate (NAR) in the first and second aerobic compartment
16
(a)
(b)
3
1.6 NH+4 loading rate
A
Maximum capacity 1.2
B
kg N/(kg VSS*d)
2
C
1.5
4
kg N-NH+ /(kg VSS*d)
1.4
NH+4 Removal
2.5
1
1 0.8 0.6
C
B
A
0.4 0.5
0
0.2
0
20
40
0
60
Time (d)
0
20
40
60
Time (d)
Fig. 3 –NH4+ loading rate, removal and maximum NH4+ removal capacity in the (a) first and (b) second aerobic compartment
17
(a)
(b)
21
3.2
20
2.8
2
19.5 19 18.5 18 17.5 17
Measured OUR Curve fit OUR
16.5 16
Measured OUR Curve fit OUR
3
Oxygen uptake rate (mg O /(L*h))
2
Oxygen uptake rate (mg O /(L*h))
20.5
0
0.1
0.2 Time (Day)
0.3
2.6 2.4 2.2 2 1.8 1.6
0.4
0
0.2
0.4 0.6 Time (Day)
0.8
1
Fig. 4 – OUR measurements used for the determination maximum specific growth rate of (a) AOB and (b) NOB
18
(a)
(b) NH+, NO- ,NO- (mg N/L)
M-NH+4 35
35
M-NO-2
S-NH+4
25
3
25
4
S-NO-2 20
S-NO-3 20
15
15
10
10
5
5
4
2
30
2
M-NO-3
3
NH+, NO- ,NO- (mg N/L)
30
0
0
10
20
30 Time (d)
40
50
0
60
0
10
20
30 Time (d)
40
50
60
Fig. 5 – Measured (M) and simulated (S) NH4+, NO2- , NO3- concentration in (a) first and (b) second aerobic compartment
19
(a)
(b)
2.5
2.5
2
2
DO (mg O N/L)
1.5
2
2
DO (mg O N/L)
NOB proliferation
1
AOB abundant 1.5 NOB proliferation 1
NOB washout with FA inhibition 0.5
0.5 AOB insufficient
NOB washout
NOB washout without FA inhibition 3
4
5
6
7
8
3
SRT (d)
4
5
6
7
8
SRT (d)
Fig. 6 – (a) NOB washout or proliferation under different SRT and DO concentration for model with or without FA inhibition on NOB; (b) SRT and DO combination (area marked with light blue) that can be used for partial nitrification operation predicted by model without FA inhibition on NOB.
20
Table 1 – Experimental plan and operational conditions Period A B C Date (d) 0-23 24-42 43-58 Recirculation ratio R1 1 1 2.5 Recirculation ratio R2 1 1 1 Influent TAN (mg N/L) 50.0± 4.6 105.0±6.8 105.0±6.8 pH in the First compartment 7.5±0.2 8.7±0.2 8.85±0.15
21
S2213-3437(16)30248-2 http://dx.doi.org/doi:10.1016/j.jece.2016.06.037 JECE 1170
To appear in: Received date: Revised date: Accepted date:
18-3-2016 7-6-2016 30-6-2016
Please cite this article as: Jun Wu, Yue Zhang, Gangyan, Differentiating two partial nitrification mechanisms: Inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity, Journal of Environmental Chemical Engineering http://dx.doi.org/10.1016/j.jece.2016.06.037 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Differentiating two partial nitrification mechanisms: inhibiting nitrite oxidizing bacteria activity or promoting ammonium oxidizing bacteria activity
Jun Wu*, Yue Zhang, Gangyan
School of Environmental Engineering and science, Yangzhou University, 196 West Huayang Road, Yangzhou, Jiangsu, 225127, China. Tel: +86-514-87971389, Fax: +86-514-87978626
*Correspondence author Email address: [email protected]
1
Highlights
Partial nitrification was closely associated to the specific biomass NH4+ load;
A low SRT was needed to maintain high specific biomass NH4+ load;
FA inhibition of NOB was insignificant for partial nitrification;
Increased AOB activity was the main reason for partial nitrification ;
Abstract Partial nitrification is usually more easily achieved for high NH4+ concentration wastewater. In this study, a two-compartment aerobic nitrification reactor was used to raise the NH4+ concentration in the first aerobic compartment and to enhance partial nitrification for low NH4+ concentration wastewater. In addition to the regular sludge wastage to maintain sludge retention time (SRT), extra sludge wastage was used to increase the specific biomass NH4+ load. The experimental results indicated that partial nitrification was closely associated to the specific biomass NH4+ load. The modelling results indicated that a low SRT was needed to maintain the low biomass concentration and high specific biomass NH4+ load. The increased AOB (ammonium oxidizing bacteria) activity, instead of the free ammonia (FA) inhibition of NOB (nitrite oxidizing bacteria) was identified to be the main mechanism for partial nitrification in low NH4+ concentration wastewater treatment with high specific biomass NH4+ load.
Keywords: nitritation; sludge retention time; ammonium load; two-compartment nitrification
2
1. Introduction In the partial nitrification (nitritation), the ammonium (NH4+) oxidation was controlled at nitrite (NO2-) step, without being further oxidized into nitrate (NO3-) [1]. The partial nitrification offers several distinctive advantages including oxygen saving in the nitrification process, reduced carbon source requirement in the denitrification process and less sludge production [2]. Partial nitrification is also a requirement for the application of the recently developed autotrophic nitrogen removal by anaerobic ammonium oxidization (ANAMMOX) [3]. Applying oxygen limitation is one of the most widely used techniques for partial nitrification [4]. The rationale behind this operation is that NOB (nitrite oxidizing bacteria) has lower DO (dissolved oxygen) affinity than AOB (ammonium oxidizing bacteria) [5, 6]. Therefore, NOB will be washed out under DO limitation. However, recent studies have indicated the certain species of NOB could have higher DO affinity than AOB [7, 8], therefore, applying low DO will cause more inhibition in AOB activity than in NOB activity. Furthermore, the effect of low DO on partial nitrification is also complicated by other NOB inhibiting factors. Free ammonia (FA) inhibition of NOB was considered to another measure or achieving partial nitrification [9]. In studies that relied on FA inhibition on NOB for partial nitrification operation, DO limitation was also applied for a combined inhibition on NOB by FA and low DO [10, 11]. It is usually difficult to differentiate the exact contribution of NOB inhibition by low DO or FA. To elucidate the exact contribution of partial nitrification by FA, the DO was supplied in non-limiting condition in this study. Partial nitrification is usually assumed to be more easily achieved for high NH4+ load condition [12]. The FA inhibition on NOB was thought to be the main reason; however, besides reducing NOB activity by FA inhibition, the contribution of high NH4+ concentration on NO2- accumulation can also be explained by the increased AOB activity due to the high Monod term value for NH4+ [13]. No studies have been carried out to elucidate the quantitative contribution of high NH4+ concentration on NO2- accumulation. Partial nitrification for low NH4+ concentration wastewater (less than ca. 100 mg N/L) is an important step to implement the recently developed ANAMMOX technique in the municipal wastewater treatment [14]. The bulk liquid NH4+ concentration in the conventional CSTR (continuously stirred tank reactor) for municipal wastewater treatment is usually too low to take advantage of the high NH4+concentration for partial nitrification. To overcome the problem, the aerobic reactor was split into two compartments. It was expected that the high NH4+ in the first compartment could enhance partial nitrification by decreasing the NOB activity (by FA inhibition), or increasing the AOB activity (by increasing the value of Monod term for NH4+). 3
Mathematical model was included to elucidate the exact mechanisms of partial nitrification in the two-compartment reactor. 2. Materials and methods 2.1. Experimental bioreactor The schematic layout of the experimental bioreactor was shown in Fig.1. It includes one anoxic compartment and two aerobic compartments (#1, #2). Each compartment has an effective volume of 2 L. The effective volume of the settlement tank is 1.5 L. The influent flow rate (Q) was maintained at 0.5 L/h, resulting hydraulic retention time (HRT) of 12 h. The recirculation flow ratio (R1 and R2) from #1 and #2 aerobic compartment to the anoxic compartment was varied according to experimental plan shown in Table-1. The sludge retention time (SRT) was adjusted by wasting 400 ml of mixed liquor from the #2 aerobic compartment. As the solids concentration in the #2 aerobic compartment was roughly twice as high as the solids concentration in the other two compartments, the effective SRT was calculated to be 10 d. Synthetic wastewater was prepared according the recipe from Wu & He [15]. The chemical oxygen demand (COD) and N-NH4+ ratio was maintained at above 6 to allow complete denitrification in the anoxic tank. The seeding sludge was taken from the aeration tank of the local wastewater treatment plant. The bioreactor was placed in an air-conditioned room with temperature controlled at 25.0±0.8 ºC. The DO concentration in both of the aerobic compartments was provided at above 6 mg O2/L to create an oxygen non-limiting condition. 2.2. Experimental plans Before taking samples for measurement, the bioreactor was operated for 20 days of assimilation period. Then the bioreactor was operated according to the experiment plan shown in Table 1. During the experiment period, the NH4+, NO2- and NO3concentration in the influent, anoxic compartment and two aerobic compartments were measured regularly. The pH and NH4+ concentration in the influent were varied to examine the effect of FA on NOB inhibition. The changes in pH, influent NH4+ concentration and recirculation ratio were also necessary to provide enough data excitation for the mathematical model calibration [16]. At the end of period A, in addition to the regular sludge wastage to maintain the SRT, an extra portion of biomass solids was wasted to further decrease the biomass concentration and increase the specific biomass NH4+ loading rate. This allowed the high NH4+ loading conditions to be investigated for low NH4+ concentration wastewater. 4
2.3. Batch experiment for the measurement of the maximum specific growth rate for AOB and NOB The oxygen uptake rate (OUR) was measured to calculate the maximum specific growth rate for AOB and NOB (μAOB and μNOB , d-1). The OUR was measured using the respirometer shown by Wu et al [17]. The OUR measured at time t under ammonium and DO non-limiting condition (OURNH(t)) can be expressed by the following equation [18]: OUR
NH
(t ) e
t(
AOB
b AOB )
OUR
0
(1)
NH
OURNH(t) is expressed by the exponential function of
OUR
0 NH
(the initial OUR after
ammonium addition). By curve fitting the measured OUR to the simulated OUR and assuming the decay rate bAOB to be 5% of μAOB , μAOB can be estimated. Similarly, the μNOB can be estimated by the following equation: OUR
NO 2
(t ) e
t(
NOB
b NOB )
OUR
0
(2)
NO 2
Where, OURNO2(t) is the OUR at time t after nitrite addition;
OUR
0 NO 2
is the initial
OUR after nitrite addition. 2.4. Sample analysis method The pH and DO were measured by HACH pH electrode and LDO oxygen sensor respectively. VSS, COD, NH4+, NO2- and NO3- concentration were measured according to the standard method [19]. 2.5. Mathematical modelling 2.5.1. Model kinetics and parameters The ASM (activated sludge model) type of model was included to simulate the reactor shown in Fig. 1 [16]. The model had 4 particulate components including AOB (XAOB), NOB(XNOB), heterotrophic bacteria (XH) and inert material (XI); and 6 soluble components including DO (SO), NH4+ (SNH), NO2- (SNO2) and NO3- (SNO3) and soluble COD (Ss). The complete model stoichiometric matrix and kinetic rate expressions were shown in the supporting information Table S1. The maximum specific growth rate for AOB and NOB were determined by respirometer shown in section 2.3. The FA inhibition constant for NOB (KIA) was determined from model calibration. Other parameter values was taken from the literature and shown in the Table S2. The growth rates AOB or NOB were expressed by the following equation: 5
dX
AOB
dt dX
NOB
dt
S AOB
K
S
S , NH
S NOB
K
SO
NH NH
K
S , NO 2
X
NO 2
K
O , NOB
(3)
AOB
SO
NO 2
S
SO
O , AOB
K SO K
IA
IA
FA
X
(4) NOB
Where, KS,NH and KS,NO2 were substrate half saturation constant for AOB and NOB; KO,AOB and KO,NOB were substrate half saturation constant for AOB and NOB; KIA was the FA inhibition constant. 2.5.2. Model calibration The NH4+, NO2- and NO3- concentration data measured during the experiment was used for model calibration. The KIA value was determined by ensuring the best curve fitting between the model prediction and experimental data. 2.4.3. Modelling strategy For the model based analysis, the influent was fixed at 50 mg N/L and pH was fixed at 7.5 to represent the municipal wastewater treatment. The simulation time was 300 days to reach steady state. The model was simulated under different combination of SRT and DO to investigate their effect on partial nitrification. To quantify the effect of FA on nitration, a new model that didn’t include the FA inhibition on NOB was run and compared to the original simulation results. In the new model, FA inhibition term for the NOB was removed. The NOB growth rate was expressed by the following equation: dX
NOB
dt
S NOB
K
SO
NO 2
S , NO 2
S
NO 2
K
O , NOB
SO
X
(5) NOB
Other kinetic rate expressions remained the same as shown in the supporting information Table S1. The AOB and NOB composition were compared for simulation results with different combination of SRT and DO. The amount of AOB was considered to be sufficient if more than 90% of NH4+ oxidation rate was achieved. The NOB washout was defined by NOB concentration of less than 0.1 mg COD/L. 3. Results and discussion 3.1. Reactor performance Fig. 2(a-c) showed the NH4+, NO2- and NO3- concentration monitored in the influent, anoxic and two aerobic compartments over the experiment periods specified in Table-1. Fig. 2(d) showed that the NAR (nitrite accumulation rate) in the two aerobic compartments, defined by the following equation:
6
NO
NAR NO
2
2
NO
(6)
100 %
3
The NO2- and NO3- concentration were near zero in the anoxic tank during the experiment periods (Fig.2(a)); therefore, a complete denitrification was achieved in the anoxic tank. During period A (day 0-23), the NH4+ was completely removed in both aerobic compartments (Fig.2(b-c)); the NO2- concentration gradually increased, whereas the NO3- concentration slightly decreased in the #1 aerobic compartment (Fig.2(b)); the NO2- was completely oxidized into NO3- in the #2 aerobic compartment (Fig.2(c)). Fig. 2(d) showed that the nitrite accmulation was low in the #1 aerobic compartment and remained at near zero in the #2 aerobic compartment during period A. In period B, the increase in influent NH4+ concentration and extra sludge solid loss of nearly 65% resulted in the high NH4+ accumulation up to 30 mg N/L in the #2 aerobic compartment (Fig. 2(b)). The measured VSS concentration in the bioreactor was shown in the Fig. S1. NO3- production was very low and nearly 100% nitrite accumulation was achieved in the #1 aerobic compartment (Fig. 2(b, d)). Despite the massive sludge loss, NH4+ accumulation in the #2 aerobic compartment was not significant (Fig. 2(c)). NO2- was initially accumulated and then decreased in the #2 aerobic compartment, with NAR increasing from ca. 60% to 100% initially and then deceasing to ca. 50% (Fig. 2(c, d)). In period C, the recirculation from the #1 aerobic reactor (R1) increased to 2.5 and the pH decreased to 8.2. The solid loss was recovered in the period C as demonstrated by the VSS measurement shown in Fig. S1. A low NO3- concentration was noticeable in the #1 aerobic compartment (Fig. 2(b)). The NO2- concentration decreased significantly in the #2 aerobic compartment (Fig. 2(c)). The NAR remained above 70% in the #1 aerobic compartment and deceased to near zero in the #2 aerobic compartment (Fig. 2(d)). Fig. 3 shows the specific biomass NH4+ loading rate and actual NH4+ removal (kg N/(kg VSS · d)) for both aerobic compartments. The specific biomass NH4+ loading rate was defined by the NH4+ loading per kg VSS per day (kg N/(kg VSS · d)). The sludge maximum NH4+ removal capacity (kg N/(kg VSS · d)) was derived from measurement of NH4+ removal at NH4+ and DO non-limiting condition in a batch reactor. During period A and C, nearly all the NH4+ load was removed in the #1 aerobic compartment. The NH4+ load entered the #2 aerobic compartment after the extra solids wastage. The maximum NH4+ removal capacity indicated that, except for the extra solid loss period, the system was operated at lower NH4+ loading rate than the NH4+ removal capacity. This explained why the NH4+ effluent concentration was not affected by the massive solid loss. 7
The high NAR (nitrite accumulation rate) was found to be closely associated with the NH4+ load. The high NAR in the first aerobic compartment could be explained by the higher NH4+ load compared to the second aerobic compartment. The sudden increasing NAR in the first aerobic compartment, following the solids loss, also pointed to the conclusion that increasing NH4+ load led to NO2- accumulation. The high NAR could not be sustained due to the recovering of biomass concentration and decreasing in the specific biomass NH4+ load. The maximum NH4+ removal capacity of ca. 1.2 kg N/(kg VSS · d) in this study was lower than the 2.87 kg N/(kg VSS · d) reported by Chen et al. [20], suggesting there was room for increasing the NH4+ load for NO2- accumulation. Because oxygen was provided in non-limiting condition, the results in Fig. 2 and Fig. 3 can suggest that NH4+ load was critical for NO2- accumulation. The result was consistent with previous reports in which NH4+ was provided in excess to promote stable partial nitrification [13]. However, these studies were carried out at extremely high NH4+ concentration (above ca. 1000 mg N/L). The result in this study indicated the partial nitrification could be achieved for low NH4+ concentration (less than ca. 100 mg N/L) by increasing the specific biomass NH4+ load via sludge wastage. The aggressive sludge wasting technique to achieve partial nitrification used by Regmi et al. [8] also supported the conclusion in this study. The NOB inhibition by FA under high NH4+ load was usually assumed to be the main mechanism for achieving partial nitrification [10]. On the other hand, AOB activity could be increased under high NH4+ load due to the increasing in Monod term for NH4+. The high AOB activity could also contribute to the NO2- accumulation. Although, the experiment result in this study showed that the high NH4+ load could promote NO2- accumulation. It is not sure whether it is due to the increased AOB activity or the decreased NOB activity. The modelling study was included to elucidate the mechanism. 3.2. Modelling results The maximum specific growth rates for AOB and NOB (μAOB, μNOB, d-1) were calibrated for the model shown in Table S1.The OUR measured under substrate non-limiting and high food to mass (F/M) ration condition used for measurement of μAOB and μNOB (d-1) was shown in Fig. 4 . Fig. 4(a) showed the OUR measured used for maximum specific AOB growth rate (μAOB, d-1) measurement under DO and NH4+ non-limiting condition. By curve fitting the OUR data to the equation 1, the μAOB value could be estimated at 0.98 d-1. Fig. 4(b) showed the OUR measured used for maximum specific NOB growth rate (μNOB, d-1) measurement under DO and NO2non-limiting condition. By curve fitting the OUR data to the equation 2, the μNOB value could be estimated at 0.62 d-1. The μAOB and μNOB value were higher than the normally used values [6], possibly due to the high NH4+ and NO2- load in the first 8
aerobic compartment. The high substrate availability will select fast growing r-strategist species [21]. 3.2.1. Description of the experimental results Fig. 5 showed the simulated and measured NH4+, NO2- and NO3- concentration in the first and second aerobic compartment. The initial AOB and NOB concentration and FA inhibition constant for NOB (KIA) were adjusted by trial and error method to produce the best fit between simulated and measured results. At day 24, the solids concentrations were reduced by 65% to represent the extra solid loss shown in Fig. S1. By choosing the KIA value at 0.2 (mg N/L), the simulation data was in agreement with the measured data. This confirmed that the validity of the model in represent the experimental results. The KIA value of 0.2 (mg N/L) was lower than reported by Jubany et al.[10]. The FA concentration between 0.1-1.0 mg N /L was suggested to be inhibitive to NOB [9]. Therefore, the KIA value was lower compared to the literatures. According to the equation 4, a low KIA value means a high FA inhibition of NOB. 3.2.2. Mechanisms of NO2- accumulation To quantify the effect of FA inhibition on NOB washout, simulations were carried out for model with and without FA inhibition term for NOB (equation 4 and 5). Fig. 6(a) showed the NOB washout or proliferation under different SRT and DO concentration. In general, low DO and SRT led to the NOB washout. The area below the curve marked with triangles represented NOB washout considering the FA inhibition of NOB. While the area below the curve marked with circles represented NOB washout without considering the FA inhibition of NOB. The area above the two curves represented the NOB proliferation. Compared to model without FA inhibition on NOB, the model with FA inhibition on NOB only predicts slight increasing in area that could be used for NOB washout (marked by grey shade in Fig. 6(a)). This suggested that the contribution of FA inhibition in NOB washout was insignificant for the bioreactor used in this study. It should be noted the KIA value of 0.2 (mg N/L) was lower than the values reported [10]. A low KIA value means strong inhibition. Even though, the contribution by FA inhibition for NOB washout was still insignificant. For KIA value of 0.95 (mg N/L) suggested by Jubany et al. [10], the simulation predicted a much smaller area of NOB washout contributed by FA inhibition (Fig. S2). Since the simulation result suggested a very small the contribution of FA inhibition on NOB washout, NO2- accumulation observed in experiment with high NH4+ load condition could be attributed to the increased AOB activity.
9
Fig. 6(b) showed that the SRT and DO combination (area marked with light blue) that could be used for partial nitrification operation predicted by model without FA inhibition on NOB. The area above the curve marked with squares represented condition where there was abundant AOB, defined by NH4+ oxidation rate of above 90%. The area below the curve marked with red circles showed NOB washout. The common area for the AOB abundant and NOB washout represented the SRT and DO combination required for partial nitrification (marked with light blue shade). For example, the SRT of 4 days and DO concentration of 1 mg O2/L at green dot represented one partial nitrification operation point. The low SRT predicted by the model corresponded to the massive solids loss in this study and aggressive sludge wastage used by Regmi et al. [8]. The low biomass concentration under low SRT led to high specific biomass NH4+ load, therefore, enhanced the AOB activity and resulted in NO2- accumulation. The model prediction also explained why the high NO2accumulation after extra solids loss was not sustainable under SRT of 10 days, since the solids concentration would recover and reduce the specific biomass NH4+ load. The 5-pointed red star represented full nitrification operation point and the blue square represented insufficient NH4+ oxidation. Partial nitrification achieved under high NH4+ load condition has been reported for high NH4+ concentration wastewater (ca. 500-1000 mg N/L) [10, 20, 22]. The partial nitrification achieved during massive solid loss in this study indicated that the specific biomass NH4+ load rate could be increased to achieve partial nitrification for low NH4+ concentration condition (less than ca. 100 mg N/L). However, due to the long SRT of 10 days used in the experiment, the high specific biomass NH4+ load rate could not be sustained after the recovery of biomass concentration. The simulation results indicated that a low SRT can be used to maintain the high specific biomass NH4+ load rate.
10
The increased AOB activity under high FA concentration has been reported for high NH4+ concentration wastewater (ca. 500 mg N/L)[20]. The experimental and modelling in this study confirmed that the FA inhibition was not significant in NOB washout; instead, it was the increased AOB activity under high NH4+ load condition that led to NOB washout in low NH4+ concentration wastewater. The findings provided a theoretical basis for using low SRT to achieve high specific biomass NH4+ load rate and partial nitrification in low NH4+ concentration wastewater. 4. Conclusions
Partial nitrification was closely associated to the specific biomass NH4+ load.
FA inhibition of NOB was insignificant in the two-compartment reactor for partial nitrification.
The increased AOB activity was identified to be the main mechanism of partial nitrification for low NH4+ concentration wastewater treatment with high specific biomass NH4+ load.
Acknowledgement The financial support from Natural Science Foundation of China (Grant number: 51478410) is highly appreciated. The authors also thank the support from the Yangzhou University high-end talent program.
11
Reference [1] O. Turk, D.S. Mavinic, Preliminary assessment of a shortcut in nitrogen removal from wastewater, Canadian Journal of Civil Engineering, 13 (1986) 600-605. [2] S. Aslan, L. Miller, M. Dahab, Ammonium oxidation via nitrite accumulation under limited oxygen concentration in sequencing batch reactors, Bioresour. Technol., 100 (2009) 659-664. [3] M.S.M. Jetten, S.J. Horn, M.C.M. van Loosdrecht, Towards a more sustainable municipal wastewater treatment system, Water Sci. Technol., 35 (1997) 171-180. [4] Y. Ma, Y. Peng, S. Wang, Z. Yuan, X. Wang, Achieving nitrogen removal via nitrite in a pilot-scale continuous pre-denitrification plant, Water Res., 43 (2009) 563-572. [5] A. Guisasola, I. Jubany, J.A. Baeza, J. Carrera, J. Lafuente, Respirometric estimation of the oxygen affinity constants for biological ammonium and nitrite oxidation, J. Chem. Technol. Biotechnol., 80 (2005) 388-396. [6] U. Wiesmann, Biological nitrogen removal from wastewater, Adv. Biochem. Eng. Biotechnol., 51 (1994) 113-154. [7] R. Manser, W. Gujer, H. Siegrist, Consequences of mass transfer effects on the kinetics of nitrifiers, Water Res., 39 (2005) 4633-4642. [8] P. Regmi, M.W. Miller, B. Holgate, R. Bunce, H. Park, K. Chandran, B. Wett, S. Murthy, C.B. Bott, Control of aeration, aerobic SRT and COD input for mainstream nitritation/denitritation, 57 (2014) 162-171. [9] A.C. Anthonisen, R.C. Loehr, T.B. Prakasam, E.G. Srinath, Inhibition of nitrification by ammonia and nitrous acid, J. Water Pollut. Control Fed., 48 (1976) 835-852. [10] I. Jubany, J. Lafuente, J.A. Baeza, J. Carrera, Total and stable washout of nitrite oxidizing bacteria from a nitrifying continuous activated sludge system using automatic control based on Oxygen Uptake Rate measurements, Water Res., 43 (2009) 2761-2772. [11] G. Ruiz, D. Jeison, R. Chamy, Nitrification with high nitrite accumulation for the treatment of wastewater with high ammonia concentration, Water Res., 37 (2003) 1371-1377. [12] D. Wei, X. Xue, L. Yan, M. Sun, G. Zhang, L. Shi, B. Du, Effect of influent ammonium concentration on the shift of full nitritation to partial nitrification in a sequencing batch reactor at ambient temperature, Chem. Eng. J., 235 (2014) 19-26.
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[13] E. Isanta, C. Reino, J. Carrera, J. Perez, Stable partial nitritation for low-strength wastewater at low temperature in an aerobic granular reactor, Water Res., 80 (2015) 149-158. [14] T. Lotti, R. Kleerebezem, Z. Hu, B. Kartal, M.K. de Kreuk, C. van Erp Taalman Kip, J. Kruit, T.L.G. Hendrickx, M.C.M. van Loosdrecht, Pilot-scale evaluation of anammox-based mainstream nitrogen removal from municipal wastewater, Environ. Technol., 36 (2015) 1167-1177. [15] J. Wu, C. He, Effect of cyclic aeration on fouling in submerged membrane bioreactor for wastewater treatment, Water Res., 46 (2012) 3507-3515. [16] M. Henze, W. Gujer, T. Mino, M. van Loosdrecht, Activated Sludge Models: ASM1, ASM2, ASM2d and ASM3 IWA Publishing, London, 2000. [17] J. Wu, C. He, The effect of settlement on wastewater carbon source availability based on respirometric and granulometric analysis, Chem. Eng. J., 189-190 (2012) 250-255. [18] A. Ramdani, P. Dold, S. Deleris, D. Lamarre, A. Gadbois, Y. Comeau, Biodegradation of the endogenous residue of activated sludge, Water Res., 44 (2012) 2179-2188. [19] AWWA, Standard Methods for the Examination of Water and Wastewater, 21st ed., American Water Works Association, 2005. [20] J. Chen, P. Zheng, Y. Yu, Q. Mahmood, C. Tang, Enrichment of high activity nitrifers to enhance partial nitrification process, Bioresour. Technol., 101 (2010) 7293-7298. [21] A. Terada, S. Sugawara, T. Yamamoto, S. Zhou, K. Koba, M. Hosomi, Physiological characteristics of predominant ammonia-oxidizing bacteria enriched from bioreactors with different influent supply regimes, Biochem. Eng. J., 79 (2013) 153-161. [22] L.-Y. Chai, M. Ali, X.-B. Min, Y.-X. Song, C.-J. Tang, H.-Y. Wang, C. Yu, Z.-H. Yang, Partial nitrification in an air-lift reactor with long-term feeding of increasing ammonium concentrations, Bioresour. Technol., 185 (2015) 134-142.
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Figure captions:
Fig.1 – Schematic layout of the experimental bioreactor Fig. 2 – (a) Influent NH4+ concentration (NH4+ #0) and NH4+, NO2- , NO3- in the anoxic tank (#a); (b) NH4+, NO2- , NO3- concentration in the first aerobic compartment (#1); (c) NH4+, NO2- , NO3- concentration in the second aerobic compartment (#2); (d) Nitrite accumulation rate (NAR) in the first and second aerobic compartment Fig. 3 –NH4+ loading rate, removal and maximum NH4+ removal capacity in the (a) first and (b) second aerobic compartment Fig. 4 – OUR measurements used for the determination maximum specific growth rate of (a) AOB and (b) NOB Fig. 5 – Measured (M) and simulated (S) NH4+, NO2- , NO3- concentration in (a) first and (b) second aerobic compartment Fig. 6 – (a) NOB washout or proliferation under different SRT and DO concentration for model with or without FA inhibition on NOB; (b) SRT and DO combination (area marked with light blue) that can be used for partial nitrification operation predicted by model without FA inhibition on NOB.
14
Figures:
Fig.1 – Schematic layout of the experimental bioreactor
15
A
-
C
40
D
0
NH+4 #1
Solids loss
NO-2 #1
30
D
NO-3 #1
20
A
B C
10
+
20 0
10
20
30 40 Time (day)
50
60
0
0
10
20
30 40 Time (day)
NH+4 #2
30
NO-2 #2
B
D
C
NO-3 #2
20 10
0
10
-
A
+
0
20
50
60
(d)
NO2 accmulation rate (%)
(c) 40
-
-
N-NH4, N-NO2, N-NO3 (mgN/L)
+
-
B
(b) 40
-
80
N-NH4, N-NO2, N-NO3 (mg N/L)
100
60
(a)
NH+4 #a NO-2 #a NO-3 #a
-
N-NH4, N-NO2, N-NO3 (mgN/L)
NH+4 #0
120
30 40 Time (day)
50
100
60
C A
40
D
B
20 0
60
Aerobic #2 Aerobic #1
80
0
10
20
30 40 Time (day)
50
60
Fig. 2 – (a) Influent NH4+ concentration (NH4+ #0) and NH4+, NO2- , NO3- in the anoxic tank (#a); (b) NH4+, NO2- , NO3- concentration in the first aerobic compartment (#1); (c) NH4+, NO2- , NO3- concentration in the second aerobic compartment (#2); (d) Nitrite accumulation rate (NAR) in the first and second aerobic compartment
16
(a)
(b)
3
1.6 NH+4 loading rate
A
Maximum capacity 1.2
B
kg N/(kg VSS*d)
2
C
1.5
4
kg N-NH+ /(kg VSS*d)
1.4
NH+4 Removal
2.5
1
1 0.8 0.6
C
B
A
0.4 0.5
0
0.2
0
20
40
0
60
Time (d)
0
20
40
60
Time (d)
Fig. 3 –NH4+ loading rate, removal and maximum NH4+ removal capacity in the (a) first and (b) second aerobic compartment
17
(a)
(b)
21
3.2
20
2.8
2
19.5 19 18.5 18 17.5 17
Measured OUR Curve fit OUR
16.5 16
Measured OUR Curve fit OUR
3
Oxygen uptake rate (mg O /(L*h))
2
Oxygen uptake rate (mg O /(L*h))
20.5
0
0.1
0.2 Time (Day)
0.3
2.6 2.4 2.2 2 1.8 1.6
0.4
0
0.2
0.4 0.6 Time (Day)
0.8
1
Fig. 4 – OUR measurements used for the determination maximum specific growth rate of (a) AOB and (b) NOB
18
(a)
(b) NH+, NO- ,NO- (mg N/L)
M-NH+4 35
35
M-NO-2
S-NH+4
25
3
25
4
S-NO-2 20
S-NO-3 20
15
15
10
10
5
5
4
2
30
2
M-NO-3
3
NH+, NO- ,NO- (mg N/L)
30
0
0
10
20
30 Time (d)
40
50
0
60
0
10
20
30 Time (d)
40
50
60
Fig. 5 – Measured (M) and simulated (S) NH4+, NO2- , NO3- concentration in (a) first and (b) second aerobic compartment
19
(a)
(b)
2.5
2.5
2
2
DO (mg O N/L)
1.5
2
2
DO (mg O N/L)
NOB proliferation
1
AOB abundant 1.5 NOB proliferation 1
NOB washout with FA inhibition 0.5
0.5 AOB insufficient
NOB washout
NOB washout without FA inhibition 3
4
5
6
7
8
3
SRT (d)
4
5
6
7
8
SRT (d)
Fig. 6 – (a) NOB washout or proliferation under different SRT and DO concentration for model with or without FA inhibition on NOB; (b) SRT and DO combination (area marked with light blue) that can be used for partial nitrification operation predicted by model without FA inhibition on NOB.
20
Table 1 – Experimental plan and operational conditions Period A B C Date (d) 0-23 24-42 43-58 Recirculation ratio R1 1 1 2.5 Recirculation ratio R2 1 1 1 Influent TAN (mg N/L) 50.0± 4.6 105.0±6.8 105.0±6.8 pH in the First compartment 7.5±0.2 8.7±0.2 8.85±0.15
21