Incorporating the Oxic-Settling-Anaerobic (OSA)

Process into an Anoxic–Oxic System for Sewage

Sludge Reduction and Nutrient Removal
Pablo Maria Romero Pareja ,a Carlos Alberto Aragon Cruz,b Jose Maria Quiroga Alonso,a and
Marıa Dolores Coello Oviedoa
a
Department of Environmental Technology, Faculty of Marine and Environmental Sciences, Universidad de Cadiz, Poligono Rio
San Pedro s/n, 11510 Puerto Real, Cadiz, Spain; pablo.romero@uca.es (for correspondence)
b
Foundation Centre for New Water Technologies (CENTA), Autovia Sevilla-Huelva (A-49) km. 28, 41820 Carrion de los Cespedes,
Seville, Spain
Published online 27 October 2017 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/ep.12784

The production of sludge in biological wastewater treat-
ments is a growing problem worldwide as it entails eco-
nomic, environmental, and health problems. The oxic-
settling-anaerobic (OSA) process is one of the techniques
that have been developed to reduce the sludge produced in
the activated sludge process (ASP). This article reports the
feasibility of the application of the OSA process in a nutrient
removal system, specifically, in an anoxic–oxic (A) process.
The results reveal TN removal in the A 1 OSA system of
68.84% and 69.91% (for 7 and 11 h of sludge anaerobic
exposure time, SAET, respectively) versus 38.02% and 32.47%
in the OSA system with similar SAET. The percentage sludge
reductions obtained were, respectively, 22.41% and 31.93%
in the OSA system and 23.53% and 32.49% in the A 1 OSA
system compared to the sludge produced in the conventional
ASP. The study confirms the suitability of the application of
the OSA process in a nutrient removal system, as nutrient
removal performances are not affected, while at the same
time achieving higher sludge reduction. V C 2017 American Insti-
tute of Chemical Engineers Environ Prog, 37: 1068–1074, 2018
Keywords: sewage sludge reduction, OSA process, biologi-
cal nutrient removal
INTRODUCTION
Efficient wastewater treatment is a key aspect in establish-
ing the effective development of a country, as it results in
progress in environmental, health, and economic terms for
the entire population. Although biological treatment is the
most widely-used core process in wastewater treatment, it
entails an important drawback, namely the production of a
large amount of sludge which needs to be managed prior to
disposal, constituting up to 65% of the total operating costs
of a conventional wastewater treatment plant (WWTP) [1]. As
a result, the development and study of strategies and techni-
ques for reducing sludge production is a field of growing
interest [1]. Moreover, minimization of excess sludge produc-
tion in the water line of the wastewater treatment process,
C 2017 American Institute of Chemical Engineers
V mining the mechanism of nutrient removal.
rather than post-treatment of the sludge which is produced,
is the preferable way to address the problem [2]. Different
techniques, known as side-stream techniques, have been
developed for this purpose. These can be classified in four
groups according to the predominant mechanism causing
sludge reduction: cell lysis-cryptic growth, endogenous
metabolism, uncoupled metabolism, and microbial predation
[3]. One of the most promising side-stream techniques is the
oxic-settling-anaerobic (OSA) process, which consists in the
addition of an anaerobic tank to the recirculation line, in
which sludge is maintained under low oxidation reduction
potential (ORP) values [1]. The reduction in sludge produced
using the oxic-settling-anaerobic (OSA) process reported in
the literature varies between 15% and 65% [4,5]. In contrast
to other techniques, this reduction is not adduced to one sin-
gle mechanism [2,4,6]. Phenomena of uncoupled metabolism,
sludge decay, reactions occurring in an anaerobic environ-
ment, release of extracellular polymeric substances, and
domination of slow-growing organisms have been identified
as the causes of sludge reduction employing the OSA pro-
cess [3,7,8]. To date, most OSA studies have been conducted
employing a single aerobic tank as bioreactor, there being
few studies in which the OSA process is employed in more
complex systems [2], specifically in those aimed at enhancing
nutrient removal [4]. In this study, two identical lab-scale
plants were operated in parallel. The first plant operated as a
conventional activated sludge process (ASP), in which the
OSA process was applied. The second consisted of a Lud-
zack–Ettinger system (anoxic/oxic biological reactor, hence-
forth A) to which the OSA process was also applied. This
setup, referred to as the A 1 OSA system, was developed by
Zhou et al. [9]. It has also been recently studied by
Khursheed et al. [3], obtaining better results in terms of
sludge reduction compared to an A system. This study
reports further details regarding nutrient removal throughout
the four operating schemes (conventional ASP, conventional
OSA, A system, and A 1 OSA). This article also presents a
detailed discussion of nutrient concentration profiles in the
different units of the A and A 1 OSA process aimed at deter-

1068 May/June 2018 Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep
Figure 1. Scheme of both plants: (a) fully aerated biological reactor 1 OSA, conventional OSA system; (b) Ludzack–Ettin-
ger 1 OSA, A 1 OSA system. [Color figure can be viewed at wileyonlinelibrary.com]

Table 1. Operative parameters and conditions of the plants during the study.

Phase Anaerobic Recirculation HRT SAET Duration, DO

of study tank rate (h) (h) days Section (mgO2/L) ri
Pilot plant operating a 0 No 1.5 9 – 60 Aerobic section 3.5–4.5 –
fully aerated process (conventional)
(Plant A) I Yes 1.5 9 7 45 Anaerobic tank –
II Yes 1.5 9 11 50
Pilot plant operating 0 No 1.5 9 – 60 Aerobic section 3.5–4.5 1.8–2.2
an anoxic–oxic (conventional)
process (Plant B) I Yes 1.5 9 7 45 Anoxic section <0.2
II Yes 0.9 9 11 40 Anaerobic tank – –

HRT, hydraulic residence time; ri, internal recirculation (Ludzack–Ettinger process)

MATERIALS AND METHODS
Pilot Plant Setup
Two similar lab-scale plants were employed in this study.
Both units were made of methacrylate. The biological reac-
tors, settling units, and anaerobic tanks (OSA process) used
in both setups had a volume of 13.2, 6, and 14 L, respec-
tively. All volumes were equipped with agitators to avoid
scumming and biofouling. The entire volume of the biologi-
cal reactor was aerated throughout the study in the reference
unit (Plant A). In the second plant (Plant B), this volume was
divided by means of a methacrylate sheet to provide two dis-
tinct areas (one anoxic and the other oxic) with the anoxic–
aerobic process system being developed to enhance nitrogen
removal.
Experimental Operation
Both lab-scale plants first operated following the scheme
of a conventional ASP (Phase 0). Subsequently, anaerobic
tanks were added to the recirculation lines of both plants to
operate in OSA mode. Figure 1 shows the setup of both
plants.
The OSA process was successively run in both plants. The
set values for the operating parameters and the duration of
the stages are shown in Table 1, together with values for
other operating parameter.
Purging of the sludge was carried out to maintain a stable
solids concentration, this waste being considered as excess
sludge produced (ESP). Moreover, the values of accumulative
sludge produced led to a solids retention time of 11.5 days,
21.7 days, and 46.1 days in pilot plant A for Phases 0, I, and
II, respectively, and of 12.2 days, 23.6 days, and 40.3 days in
Plant B for Phases 0, I, and II, respectively.
The operating parameters of the plants were controlled
by means of a PLC (Mod. SR2B202B, Schneider Electric,
Rueil-Malmaison, France) programmed from a PC. Phase II
(sludge anaerobic exposure time, SAET: 11h) lasted 10 days
less in Plant B than in Plant A, as there was a leakage caused
by breakage of a valve in the anaerobic tank on day 38 that
led to a major mass loss and resulted in the study being
interrupted.
Experimental Wastewater
The influent to the plants comprised 90% synthetic water
(SW) and 10% actual wastewater taken from an urban
WWTP near to the research center (Jerez de la Frontera
WWTP). The SW was made up according to Ye et al. [8],
though diluting to double volume and maintaining a final
concentration of KH2PO4 and urea of 30 and 25 mg/L,
respectively, so as to obtain an influent with appreciable
phosphorous and ammonia concentrations and thus highlight
the performance of the nutrient removal processes, as was
done in similar studies [5]. The room temperature ranged
between 18 and 258C throughout the study.
The average values of the characteristics of the influent
are summarized in Table 2.
Analysis and Calculation
Values of total suspended solids (TSS), VSS, MLSS, mixed
liquor suspended (volatile) solids (MLVSS), sludge volume
index (SVI), and COD were measured 4 days a week accord-
ing to the Standard Methods [10]. MLSS and MLVSS were
determined in the sludge from the biological reactors of both
plants, the anaerobic tanks of both plants (only during OSA
stages), and waste sludge. TSS, VSS, and COD values were
determined in the influent and effluents of both plants. SVI

Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep May/June 2018 1069
Table 2. Average influent characteristics.
Parameter
COD, mg O2/L 325.5 6 18.4
Total nitrogen, mg N/L 53.1 6 8.4
Ammonia nitrogen, mg NH4-N/L 38.1 6 7.3
SS, mg/L 21.1 6 6.2
Total phosphorous, mg P/L 12.5 6 0.7
was determined using sludge from the biological reactors of
both plants. ORP and dissolved oxygen (DO) values were
measured using the corresponding probes: ORP meter PCE-
228 and WTWT oxi330i/SET.
All nutrient (total phosphorous (TP), PO32 2
4 , NO3 , and
1
NH4 ) analyses were conducted 2–3 times a week using
Merck kits (Merck, Darmstadt, Germany) for the influents
and effluents of both plants. Total nitrogen (TN) was deter-
mined on a Shimadzu TOC-LCPH equipped with a TN mea-
surement unit (Shimadzu, Kyoto, Japan). Sludge samples
were taken 1–2 times a week from the anoxic and aerobic
sections and the anaerobic tank (during the OSA phases) in
Plant B to measure ammonia, nitrate, and phosphate concen-
trations in solution. For this purpose, the sampled sludge
from each tank was filtered through a 0.45 mm fiber filter
(Pall Corporation, Port Washington, NY).
The observed sludge yield (Yobs) was used in this study to
express sludge generation. The general equation for Yobs is:
Cm
Yobs 5 ; The average TSS values only increased above 15 mg/L
CS
where Cm is the cumulative produced sludge (g MLSS) and
CS is the cumulative consumed substrate.
The overall Yobs was obtained for every phase of both
plants as the slope of the regression line of the chart represent-
ing Cm,t versus CS,t for the period comprising every phase.
Cumulative produced sludge was calculated by consider-
ing the variation in solids concentration in the biological
reactors and anaerobic tanks (OSA stages), cumulative sludge
from purged waste sludge (including sludge samples), and
solids loss in the effluent. It is expressed in following equa-
tion, adaptation of Chon et al. [11]:
Cm;t 5Van  DXan 1Vre  DXre 1 Vp  Xp 1Qe  Xe  t1Vans  Xan 1Vres  Xre
where Cm,t is the cumulative produced sludge for day t, Van is
the volume of the anaerobic tank (L), Vre is the volume of the
biological reactor (L), Qe is the effluent daily flow rate (Ld21),
Xe is the TSS of the effluent (gL21), t is the time from the previ-
ous measurement of Cm (d), DXan is the change in value of
MLSS in the anaerobic tank since the previous measurement of
Cm (gL21), DXre is the change in value of MLSS in the biologi-
cal reactor since the previous measurement of Cm (gL21), Vp is
the volume of purged sludge on day t (L), Xp is the MLSS of
purged sludge(gL21), Vans is the volume of sampled sludge
from the anaerobic tank on day t (L), Xan is the MLSS of sludge
in the anaerobic tank (gL21), Vres is the volume of sampled
sludge from the biological tank on day t (L), and Xre is the
MLSS of sludge in the biological reactor (gL21).
The cumulative consumed substrate was calculated
according to the following expression:
X
t
Cs;t 5 ððSin  Qin 2Se  Qe Þ  t Þ;
0
where Cs,t is the cumulative consumed substrate for day t, Sin
is the substrate concentration in the influent (g sCODL21),
Qin is the daily flow rate entering the plant (Ld21), and Se is
the substrate concentration in the effluent (g sCODL21).
1070 May/June 2018 Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep
RESULTS AND DISCUSSION
General Performances of the Processes
The solids concentrations in the biological reactor and the
anaerobic tank (OSA process only) were kept around 3.0
and 5.5 g/L, respectively, throughout the study (see the Sup-
porting Information for the evolution of MLSS in both
plants). Specifically, the MLSS concentrations were, respec-
tively, 3.06 g/L 6 0.27 and 2.96 g/L 6 0.21 in the biological
reactor in Plants A and B and 5.42 g/L 6 0.34 and 5.09 g/
L 6 0.51 in the anaerobic tank during the OSA stages in
Plants A and B.
Table 3 shows the average values of the main perfor-
mance parameters and ORP values in the anaerobic tank for
both plants.
The evolution of the sludge settling property showed a
similar trend in Plants A and B, the results confirming that
the sludge settling property is improved by the incorporation
of the anaerobic tank. This improvement was slightly lower
when operating at an SAET of 11 h (Phase II). Ye et al. [8]
found poorer settling properties when the OSA system oper-
ated at high SAET. However, during Phase II in Plant B, this
finding could also be related to the lower recirculation rate
(r) imposed (only 0.9), which prolonged the settling time. In
general, the improvement in settling properties in the OSA
system is related to the release of intracellular matter during
anaerobic exposure which exerts a floc binding effect [8,12]
and also to the proliferation of phosphate accumulative
organisms (PAO), as these form dense clusters [5].
during the conventional phase in Plant B. This circumstance
is related to occasional solid losses in the settler as a conse-
quence of the poor settling property, a fact revealed by the
very high SVI during this phase (227.20 6 23.21 mL/mgTSS).
The solids contents in the effluent were lower when the sys-
tem ran in OSA mode in both plants. Subsequently, the aver-
age solids content in the effluent from Plant A increased
when the system shifted from Phase I to Phase II. When the
same change was carried out in Plant B, however, the solids
content in the effluent decreased. In all the OSA stages, efflu-
ent TSS values were below the legal limit of 15 mgTSS/L (EU
Directive 91/271). The improvement in SS removal in the
A 1 OSA system was likewise confirmed by Zhou et al. [9].
Although the percentage COD removal was always above
the minimum legal limit of 75% COD removal (EU Directive
91/271), a negative trend was observed during the OSA
phases in Plant B (see the Supporting Information for COD
removal throughout study). No important differences were
found between the percentage COD removal during the two
conventional processes (Phase 0, Plants A and B) or between
different OSA phases in the plants. A slight improvement in
COD removal was observed in Plant A following the addition
of the anaerobic tank. This has been a common finding in
the literature [12] and may well be a consequence of the
higher substrate removal capacity of microorganisms starved
in the anaerobic tank [1,6]. Nonetheless, the observed differ-
ences in COD removal throughout the study are almost neg-
ligible and the release of COD which occurred in the
anaerobic tank at low ORP values during the OSA stages
[12,13] may be considered to have a minimal impact on over-
all performance [2,14].
Reduction in Excess Sludge Production
The plot of cumulative produced sludge versus cumula-
tive consumed substrate in the two plants throughout the
study is shown in Figure 2, where the overall Yobs for each
phase was obtained from the slope of the respective regres-
sion line.
Table 3. Main performance parameters and ORP in anaerobic tank of both plants during the study.

Phase ORP, avg. TSS effluent, avg. COD removal, avg. SVI, avg.
Plant of study value (mV) value 6 SD (mgTSS/L) value 6 SD (%) value 6 SD (mL/mgTSS)
A 0 – 13.74 6 2.84 89.70 6 4.84 160.15 6 12.10
I 2232.80 8.15 6 4.17 92.74 6 5.17 95.54 6 16.87
II 2257.99 12.62 6 1.62 91.15 6 3.62 111.04 6 5.12
B 0 – 15.88 6 1.98 90.47 6 2.88 227.20 6 23.21
I 2322.90 10.61 6 2.26 88.61 6 5.26 143.36 6 29.79
II 2311.87 9.13 6 0.79 87.66 6 3.79 152.06 6 9.12

A reduction in ESP between Phase 0 and the OSA phases

can be observed for both plants. Specifically, the percentage
reductions were 22.41% (Phase I) and 31.93% (Phase II) in
Plant A and of 23.53% (Phase I) and 32.49% (Phase I) in
Plant B. Compared with the Yobs of the conventional A pro-
cess (Phase 0, Plant B), the reductions in observed growth
rates obtained in Plant B are 16.77% (Phase I) and 26.52%
(Phase II). These lower relative sludge reduction rates
observed in the OSA stages are due to the lower growth
yield of the conventional stage of Plant B, where anoxic
reactions with an associated lower biomass growth yield are
involved [1,15].
The percentage reductions in ESP obtained by Khursheed
et al. [3] in a similar A 1 OSA system are close to those of
this study. Specifically, the aforementioned authors obtained
a reduction in ESP of between 14.6% and 39.8%. The results
obtained by Zhou et al. [9] pointed to a higher sludge reduc-
tion in ESP in an A 1 OSA system of up to 32%.

Nutrient Removal

Nitrate, Ammonia and Phosphate Concentration in the Effluent

Table 4 discloses average values and standard deviation
of nutrient concentration in effluent during the three phases
and both plants. The OSA process led to a reduction in
nitrate concentration in the effluent from both plants. Specifi-
cally, nitrate concentrations in the effluent from Plant A were
19.46 6 4.83 mg NO3-N/L, 10.05 6 2.77 mg NO3-N/L, and
17.00 6 3.87 mg NO3-N/L for Phases 0, I, and II, respectively.
The values in Plant B were 9.56 6 2.54 mg NO3-N/L, 7.14 6
1.41 mg NO3-N/L, and 5.13 6 0.76 mg NO3-N/L for consecu-
tive phases. This reduction has also been observed by other
authors [12,14]. Moreover, as can be seen from the reported
values, the nitrate concentration in the effluent from Plant B
was always lower than in Plant A, as the inclusion of an
anoxic step in the biological process led to partial consump-
tion of nitrate as an electron acceptor for the degradation of
the substrate [16]. The difference in nitrate concentration
between the plants during the OSA phases was especially
noticeable for Phase II. However, the main reason for this
difference is the variation in r imposed during this period in
both plants (Table 1). A lower r (Plant B) leads to a shorten-
ing of the aerobic step compared to the same phase in Plant
A, where part of the recycled sludge was reintroduced into
the biological reactor, thereby resulting in an increase in
nitrate concentration in the sludge flowing into the settler
unit.
In general, the performances of both lab-scale plants
were satisfactory in terms of ammonia removal during all
phases of the study, except for the conventional stage in
Plant A (Phase 0), where the ammonia concentration occa-
sionally increased above 10 mg NH4-N/L. The average
ammonia concentrations in the effluent from Plant A were
9.91 6 1.12 mg NH4-N/L, 7.80 6 1.54 mg NH4-N/L and 7.15 6
0.96 mg NH4-N/L, and in Plant B, 6.04 6 1.70 mg NH4-N/L,
3.15 6 0.86 mg NH4-N/L and 2.86 6 0.83 mg NH4-N/L, for
Phases 0, I, and II, respectively. Prolongation of anaerobic
Figure 2. Observed sludge yield throughout the study in
both plants.
exposure (SAET) during the OSA phases resulted in more
efficient ammonia removal in both plants as an indirect con-
sequence of the enhancement of the denitrification process
in the anaerobic tank. Zhou et al. [9] reported that a slight
decrease in nitrification activity occurred when running an
OSA process as a result of autotrophic decay in the anaero-
bic tank. However, other authors have reported excellent
nitrification in OSA systems [1], where solids retention times
are often high enough to allow the nitrification process. Ye
et al. [8] reported similar effluent ammonia concentrations for
conventional and OSA processes (Table 4).
The lowest phosphate concentration in the effluent from
Plant A was recorded during the conventional process com-
pared to the phases during which the plant operated in OSA
mode. Specifically, phosphate concentrations in the effluent
from Plant A were 5.12 6 0.12 mg PO4-P/L, 6.27 6 0.78 mg
PO4-P/L, and 6.97 6 0.55 mg PO4-P/L for Phases 0, I, and II,
respectively. An increase in the concentration of phosphate
released into the effluent has been previously reported in

Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep May/June 2018 1071
 32
Table 4. Average NO2 1
3 , NH4 , and PO4 concentrations and standard deviations in the effluent from both plants throughout
the study.
Phase NO3- conc. effluent,
Plant of study avg. value 6 SD (mg NO3-N/L)
A 0 19.46 6 4.83
I 10.05 6 2.77
II 17.00 6 3.87
B 0 9.56 6 2.54
I 7.14 6 1.41
II 5.13 6 0.76
Figure 3. Evolution of TN and TP removal throughout the
study in both plants.
the OSA process [2,12]. Phosphate concentration values in
Plant B were 4.72 6 0.14 mg PO4-P/L, 4.72 6 0.31 mg PO4-P/
L, and 4.07 6 0.33 mg PO4-P/L for correlative phases. All
effluents from Plant B showed lower phosphate concentra-
tions, although more pronounced differences between the
two plants are noticeable for the OSA stages (Phases I and
II), especially during the A 1 OSA phase employing an 11 h
SAET (Phase II). These results evidence an improvement in
process performance when applying A 1 OSA. Nonetheless,
a phosphorous recovery stage would be necessary in any
plant applying sludge reduction strategies, such as the OSA
process, to avoid collapse of the P removal process [1], as
phosphorous can be only removed through excess sludge.
Total Nitrogen and Phosphorous Removal
The total nitrogen and phosphorous removal rates
obtained throughout the study are shown in Figure 3.
The average TN removal rate and standard deviation dur-
ing Phase 0 in Plant A was 16.19% (61.29). This rate
increased to an average value of 38.02% (61.53) for Phase I
1072 May/June 2018 Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep
NH14 conc. effluent, avg. PO34 - conc. effluent, avg.
value 6 SD (mg NH4-N/L) value 6 SD (mg PO4-P/L)
9.91 6 1.12 5.12 6 0.12
7.80 6 1.54 6.27 6 0.78
7.15 6 0.96 6.97 6 0.55
6.04 6 1.70 4.72 6 0.14
3.15 6 0.86 4.72 6 0.31
2.86 6 0.83 4.07 6 0.33
after the anaerobic tank was added to the recirculation line.
The TN removal rate decreased slightly to 32.47% (62.93)
when the SAET was set at 11 h (Phase II). This poorer TN
removal rate could be related to an incomplete denitrification
process during this phase. The sludge decay which occurred
in the anaerobic tank and the subsequent contribution as an
endogenous carbon source resulted in a more complete
denitrification process in the OSA stages [13,16,17]. The per-
centage of TN removed in Plant B was much higher than in
Plant A, particularly when the system operated in conven-
tional mode (Phase 0), increasing from 16.19% to 59.58%
(63.71). The TN removal rates obtained during the A 1 OSA
phases in Plant B were similar: 68.84% (63.15) and 69.91%
(64.99) for Phases I and II, respectively. This improvement
in TN removal in the A 1 OSA system was also found by
Zhou et al. [9], who reported a 13.4% increase in the TN
removal rate after the inclusion of an anaerobic tank.
In Plant A, the conventional process showed an average
TP removal rate and standard deviation of 15.20% (61.05).
When the anaerobic tank was inserted in the recirculation
line, the TP removal rate increased to 27.30% (61.93) and
35.39% (61.89) for Phases I and II, respectively. Wang et al.
[18] reported an improvement in the TP removal rate after
the insertion of an anaerobic tank, leading to an increase
from 11% in the conventional ASP to 53.84%. An (Ph.D. The-
sis from HKUST, 2004) also found a marked improvement in
TP removal (from 5% to 45% in the OSA system). However,
Ye et al. [8] reported only a minor improvement in the TP
removal rate (from 48.9% to 48.0% and 58.0%). The TP
removal rates for Plant B were 21.57% (61.52), 42.32%
(63.15), and 48.63% (63.90), evidencing that the inclusion
of the anaerobic stage improves the TP removal process of
the A system. A similar improvement in TP removal was also
found in a similar system [3], with an increase in percentage
phosphorous removal from 40.1% for the A system operating
alone to 76.1% in the A 1 OSA system. However, Zhou et al.
[17] reported poorer TP removal efficiency in the A 1 OSA
system, decreasing from 43.9% to 33.6%. The measured val-
ues of ORP in the anaerobic tank are similar to those of
enhanced biological phosphorous removal (EBPR) and facili-
tate the proliferation of PAO [19]. In fact, the above study
and that of Wang et al. [5] reported a notable percentage of
PAO in biomass of OSA systems. Furthermore, small varia-
tions in internal recirculation in the biological reactor in Plant
B resulted in an improvement in the TP removal rate for
Phase II, indicating that the reserves of PAO might not be
completely restored and that TP removal could be enhanced,
as observed in Yang et al. [20]. Alternating the anoxic–aero-
bic environments in Plant B during the A 1 OSA phases
enhances the TP removal observed in the OSA process.
Nutrient Concentration in Sludge from the A 1 OSA
System (Plant B)
The results of phosphate, ammonia, and nitrate concen-
tration measurements carried out on the sludge extracted
during the three stages from Plant B are shown in Figure 4.

Figure 4. Phosphate (a), nitrate (b), and ammonia (c) con-
centrations in solution in sludge from the anaerobic tank,
anoxic section, and aerobic section in plant B (conventional
A and A 1 OSA systems).
The average phosphate concentrations in this study were
16.34 and 20.44 mg PO4-P/L in the filtrate sludge in the
anaerobic tank for the system operating the A 1 OSA process
employing a 7 h and 11 h SAET (Phases I and II), respec-
tively. From these data an overall phosphate release rates in
anaerobic tank were obtained of 0.40 and 0.37 mg PO4-P/(g
MLVSSh) for Phases I and II, respectively. The increase in
anaerobic exposure time had a direct influence on the
release of phosphate [1,5,15]. No significant difference was
found in phosphate concentration for sludge from the anoxic
and aerobic sections during Phases 0 and I, being, respec-
tively, 7.64 and 7.35 mg PO4-P/L in the anoxic section, and
4.51 and 4.06 mg PO4-P/L in the aerobic section. Phosphate
concentration during Phase II was found to be 5.87 and
2.94 mg PO4-P/L in solution in the sludge from the anoxic
and aerobic section, respectively. This decrease in phosphate
concentration in the anoxic section, a consequence of a
more intense phosphate uptake, might be due, firstly, to the
more extended internal recirculation between sections of the
Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep
bioreactor as a consequence of the smaller r imposed, to the
proliferation of denitrifying PAO [15], and to the presence of
additional carbonaceous substrate in the sludge recycled
from the anaerobic tank, thereby increasing the C/P ratio.
As expected, minimum ammonia concentrations were
determined for all phases in the aerobic section: 5.98 mg
NH4-N/L (61.22), 2.98 mg NH4-N/L (60.94), and 2.54 mg
NH4-N/L (6 0.48) for Phases 0, I, and II, respectively. The
ammonium concentration in sludge from the anoxic section
during the conventional stage (Phase 0) was found to be
11.15 mg NH4-N/L (61.23). As can be observed, the maxi-
mum average ammonia concentrations during the two
A 1 OSA sub-stages were found in sludge taken from differ-
ent tanks. For Phase II, the maximum ammonia concentra-
tion (5.14 mg NH4-N/L 6 0.39) was measured in sludge from
the anaerobic tank versus 3.49 mg NH4-N/L (60.76) in
sludge taken from the anoxic section. In contrast, the maxi-
mum ammonia concentration for Phase I was found in
sludge from the anoxic section of the biological reactor
(7.81 mg NH4-N/L 6 1.46), higher than the value of 6.78 mg
NH4-N/L (60.54) obtained in sludge from the anaerobic
tank. High ammonia concentrations in the anaerobic tank
were the result of sludge decay and ammonification, which
occurred under low ORP values and starvation conditions
[13,17] and constitutes a reason for the enhancement of nitro-
gen removal in the A 1 OSA system, as nitrogenous com-
pounds should be assimilated by the biomass in further
stages of the process. Specifically, the overall ammonia
release rates for Phases I and II in anaerobic tank were,
respectively, of 0.12 and 0.05 mg NH4-N/(g MLVSSh)
According to Novak et al. [7], the increase in ammonia con-
centration is mainly caused by degradation of protein under
anaerobic conditions, which is enhanced by the solubiliztion
of iron-bound organic matter. The above finding regarding
the section with the maximum ammonium concentration for
Phase I and Phase II may point to acute enhancement of the
nitrification process in the system during Phase II. The rea-
soning is that, even though the ammonium concentrations in
the anaerobic tank were similar for both OSA phases, i.e.
6.78 mg NH4-N/L and 5.14 mg NH4-N/L for Phases I and II,
respectively, the recorded ammonium concentration in the
anoxic stage, where the influent loaded the system, is lower
for Phase II. Consequently, the denitrification and nitrifica-
tion rate in this system seems to be especially high, in line
with Paul and Liu [1], leading to the more extended TN
removal rate obtained in this study.
The nitrate concentration in the anaerobic tank (A 1 OSA
phases) was nearly zero, as reported by Wang et al. [5]. Spe-
cifically, the average nitrate concentrations for Phases I and
II were 0.80 mg NO3-N/L (60.13) and 0.28 mg NO3-N/L
(60.09), respectively. A very low nitrate concentration is the
cause of the anaerobic conditions achieved in the plant with
an ORP level below 2200 mV [6]. Moreover, as expected,
the highest nitrate concentrations were found in the aerobic
section during all three phases. During conventional running
(Phase 0) of the A process, average values of measured
nitrate concentration were 15.42 mg NO3-N/L (62.49) and
7.28 mg NO3-N/L (61.46) in solution from the aerobic sec-
tion and anoxic section, respectively. This difference eviden-
ces the achievement of the denitrification process in the
anoxic area of the A system, leading to the TN removal
obtained in the system. Similarly, the average nitrate concen-
trations for Phases I and II were, respectively, 7.49 mg NO3-
N/L (61.04) in the aerobic section and 3.22 mg NO3-N/L
(60.57) in the anoxic section and 6.49 mg NO3-N/L (60.69)
in the aerobic section and 3.78 mg NO3-N/L (61.15) in the
anoxic section. An important finding worth highlighting here
is that, although the lowest ammonium concentrations in
solution were always found in the sludge sampled from
Phase II, the nitrate concentration in both the anoxic and
May/June 2018 1073
aerobic sections during this sub-stage was of the same order
as that obtained during Phase I. This finding could point to a
confirmation of the enhancement of the nitrification and
denitrification process in this phase. Furthermore, Huang
et al. [21] reported that insertion of an extra anaerobic stage
giving rise to an A2O process might promote denitrifying
PAO, which could in turn enhance the denitrification process
by using internal energy storage (PHA) as a carbon source
instead of exogenous carbon. Coma et al. [15] argued like-
wise to explain the improvement in the phosphorous and
nitrogen removal rates obtained in a modified UCT setup.
CONCLUSIONS
The A 1 OSA process resulted in a reduction in sludge pro-
duction compared to a conventional system (23.53% and
32.49%) and to a Ludzack–Ettinger process (16.77% and 26.52%).
The sludge reduction rates achieved in the A 1 OSA sys-
tem were higher than in the OSA system.
The overall performances of the systems are not impaired
by the inclusion of anaerobic tanks in any system.
Nutrient removal is improved in both systems when
applying the OSA process, achieving enhanced removal per-
formances in the A 1 OSA system.
ACKNOWLEDGMENTS
The authors acknowledge the financial support provided
by the Agencia de Innovaci on y Desarrollo de la Junta de
Andalucıa, Spain (Proyecto Excelencia No. P09-TEP-4539
financed by ERDF). The authors wish to thank the profes-
sionalism of Mr. Paul Barnes (English revision).
LITERATURE CITED
1. Paul, E., & Liu, Y. (2012). Biological sludge minimization
and biomaterials/bioenergy recovery technologies (1st
Edition), Hoboken, NJ: John Wiley & Sons.
2. Semblante, G.U., Hai, F.I., Ngo, H.H., Guo, W., You, S.J., Price,
W.E., & Nghiem, L.D. (2014). Sludge cycling between aerobic,
anoxic and anaerobic regimes to reduce sludge production
during wastewater treatment: Performance, mechanisms, and
implications, Bioresource Technology, 155, 395–409.
3. Khursheed, A., Sharma, M.K., Tyagi, V.K., Khan, A.A., &
Kazmi, A.A. (2015). Specific oxygen uptake rate gradient
– Another possible cause of excess sludge reduction in
oxic-settling-anaerobic (OSA) process, Chemical Engi-
neering Journal, 281, 613–622.
4. Yuan, Q., Wang, H.Y., Zhou, Y.X., & Zhao, H. (2013).
The research progress on oxic-settling-anaerobic sludge
reduction process, Advanced Materials Research, 838–
841, 2726–2734.
5. Wang, J., Li, S.Y., Jiang, F., Wu, K., Liu, G.L., Lu, H., &
Chen, G.H. (2015). A modified oxic-settling-anaerobic
activated sludge process using gravity thickening for
excess sludge reduction, Scientific Reports, 5, 13972.
6. Ferrentino, R., Langone, M., Merzari, F., Tramonte, L., &
Andrettola, G. (2016). A review of anaerobic side-stream
reactor for excess sludge reduction: Configurations,
mechanisms, and efficiency, Critical Review in Environ-
mental Science and Technology, 46, 382–405.
7. Novak, J., Chon, D.H., Curtis, B.A., & Doyle, M. (2007).
Biological solids reduction using the cannibal process,
Water Environment Research, 79, 2380–2386.
8. Ye, F.-X., Zhu, R.-F., & Li, Y. (2008). Effect of sludge
retention time in sludge holding tank on excess sludge
1074 May/June 2018 Environmental Progress & Sustainable Energy (Vol.37, No.3) DOI 10.1002/ep
production in the oxic-settling-anoxic (OSA) activated
sludge process, Journal of Chemical Technology and Bio-
technology, 83, 109–114.
9. Zhou, Z., Qiao, W., Xing, C., Jiang, L.M., Gu, Y., & Wang,
L. (2015). Characterization of dissolved organic matter in
the anoxic–oxic-settling-anaerobic sludge reduction pro-
cess, Chemical Engineering Journal, 259, 357–363.
10. American Public Health Association (APHA), American
Water Works Association (AWWA) and Water Environ-
ment Federation (WEF). (2005). Standard methods for the
examination of water and wastewater (21st Edition),
Washington, DC: APHA.
11. Chon, D.H., McNamara, R., Mo Kim, Y., Young Park, K.,
& Park, C. (2011). Investigation on the sludge reduction
mechanism in the anaerobic side-stream reactor process
using several control biological wastewater treatment
processes, Water Research, 45, 6021–6029.
12. Saby, S., Djafer, M., & Chen, G.H. (2003). Effect of low
ORP in anoxic sludge zone on excess sludge production
in oxic-settling-anoxic activated sludge process, Water
Research, 37, 11–20.
13. Foladori, P., Velho, V.F., Costa, R.H.R., Bruni, L.,
Quaranta, A., & Andreottola, G. (2015). Concerning the
role of cell lysis-cryptic growth in anaerobic side-stream
reactors: The single-cell analysis of viable, dead and
lysed bacteria, Water Research, 74, 132–142.
14. Velho, V.F., Foladori, P., Andreottola, G., & Costa, R.H.R.
(2016). Anaerobic side-stream reactor for excess sludge
reduction: 5-year management of a full-scale plant, Jour-
nal of Environmental Management, 177, 223–230.
15. Coma, M., Rovira, S., Canals, J., & Colprim, J. (2015).
Integrated side-stream reactor for biological nutrient
removal and minimization of sludge production, Water
Science & Technology, 71, 1056–1064.
16. Semblante, G.U., Hai, F.I., Bustamante, H., Guevara, N.,
Price, W.E., & Nghiem, L.D. (2016). Biosolids reduction
by the oxic-settling-anoxic process: Impact of sludge
interchange rate, Bioresource Technology, 210, 167–173.
17. Zhou, Z., Qiao, W., Xing, C., An, Y., Shen, X., Ren, W.,
Jiang, L.M., & Wang, L. (2015). Microbial community
structure of anoxic–oxic-settling-anaerobic sludge reduc-
tion process revealed by 454-pyrosequencing, Chemical
Engineering Journal, 266, 249–257.
18. Wang, J.F., Jin, W.B., Zhao, Q.L., Liu, Z.G., & Lin, J.K.
(2007). Performance of treating wastewater and anti-
shockloading in oxic-settling-anaerobic (OSA) process for
minimization of excess sludge, Huanjing Kexue/Environ-
mental Science, 28, 2488–2493.
19. Sun, L., Chen, J., Wei, X., Guo, W., Lin, M., & Yu, X.
(2016). Study of the diversity of microbial communities in
a sequencing batch reactor oxic–settling–anaerobic pro-
cess and its modified process, Canadian Journal of Micro-
biology, 62, 411–421.
20. Yang, S., Guo, W., Wu, Q., Luo, H., Peng, S., Zheng, H.,
Feng, X., & Ren, N. (2015). Evaluation of in-situ sludge
reduction and enhanced nutrient removal in an inte-
grated repeatedly coupling aerobic and anaerobic and
oxic-setting-anaerobic system, Journal of Harbin Institute
of Technology (New Series), 22, 17–24.
21. Huang, P., Li, L., Kotay, S.M., Goel, R. (2014). Carbon
mass balance and microbial ecology in a laboratory scale
reactor achieving simultaneous sludge reduction and
nutrient removal, Water Research, 53, 153–167.