Effect of hydraulic retention time on nitrogen removal in domestic wastewater by partial nitritation and anammox processes

Journal of Science and Technology in Civil Engineering NUCE 2020. 14 (2): 127–136

EFFECT OF HYDRAULIC RETENTION TIME ON
NITROGEN REMOVAL IN DOMESTIC WASTEWATER BY
PARTIAL NITRITATION AND ANAMMOX PROCESSES
Nguyen Thi My Hanha,b,∗, Tran Thi Hien Hoaa
a

Faculty of Environmental Engineering, National University of Civil and Engineering,
55 Gai Phong road, Hai Ba Trung district, Hanoi, Vietnam
b
Faculty of Infrastructure Engineering and Urban Environment, Hanoi Architectural University,
Km 10, Nguyen Trai street, Thanh Xuan district, Hanoi, Vietnam
Article history:
Received 30/12/2019, Revised 19/03/2020, Accepted 22/3/2020
Abstract
The nitrogen treatment technology using the Anammox process is known to have advantages over conventional
technology of nitrification - denitrification. For the purpose of evaluating the effect of hydraulic retention time
to nitrogen removal in domestic wastewater by Anammox process, the authors conducted the study on partial
nitritation and Anammox reactors, separately. Partial nitritation (PN) reactor used Felibendy plate material
with Nitrosomonas bacteria while Anammox (AX) reactor used Felibendy cubes carrier material with strain
Candidatus Brocadia anammoxidans. This study was implemented during 210 days. The nitrogen treatment
efficiency of the system was evaluated with different hydraulic retention times (HRTs). The short HRT of
4.5 hours in the AX reactor affected to the total nitrogen treatment efficiency is low of 52.76 ± 1.29%. With
the hydraulic retention times in PN + AX reactors of 9 and 6 hours, the effluent quality met the requirements
of B-column according to QCVN 14:2008/BTNMT or QCVN 40:2011/BTNMT.
Keywords: Nitrosomonas; Candidatus Brocadia anammoxidans; partial nitritation process; Anammox process;
nitrogen treatment.
/>
c 2020 National University of Civil Engineering

1. Introduction
With the socio-economic development, the amount of domestic wastewater discharged into water
bodies is increasing and creating challenges to the environment. The main components of domestic
wastewater are suspended solids, organic substances, nutrients and microorganisms. This untreated
wastewater will cause secondary pollution for the receiving water source or water quality declination.
Nitrogen compounds are some of quality control components in National Technical Regulations on
natural water source, receiving source and discharge. According to QCVN 14:2008/BTNMT for domestic wastewater [1] or QCVN 40:2011/BTNMT for industry wastewater [2], before discharging into
the receiving bodies, which serve as sources for domestic and none-domestic water supply purposes,
total nitrogen concentrations must be less than 20 mg N/l and 40mg/l for A-column and B-column,
respectively.
∗

Corresponding author. E-mail address: (Hanh, N. T. M.)

127


Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

For the treatment of nitrogen compounds in wastewater, centralized wastewater treatment plants
use conventional biological treatment methods (aerobic); Advanced biological treatment (nitrogen
compounds and phosphorus compounds treatment). With the wastewater treatment technologies being
applied in Vietnam, some technologies can not fully handle nitrogen such as trickling biofilter (TF)
or conventional activated sludge (CAS) technology. Besides, some other technologies require internal
sludge recirculation, or require large amounts of oxygen, for example anoxic oxic (AO), anaerobic
– anoxic – oxic (A2O), sequencing batch reactor (SBR) or additional carbon sources. Applying a
different processing technology to overcome the above weaknesses is very necessary.
The discovery of anammox bacteria led to the development of a fully autotrophic process that
does not required chemical and uses less energy for aeration or mixing, offering the plants [3]. The
technology of nitrogen treatment by Anammox process need firstly, partial nitritation (partial oxidation of ammonium to nitrite, Eq. (1)) and secondly, the anammox process (anoxic combination of

ammonium and nitrite to form dinitrogen gas, Eq. (2)) [4].
NH+4 + 0.83O2 → 0.45NH+4 + 0.55NO−2 + 0.55H2 O + 1.1H+

(1)

NH+4 + 1.32NO2 + 0.066HCO−3 + 0.13H+ → 1.02N2 + 0.26NO−3 + 0.066CH2 O0.5 N0.15 + 2.03H2 O (2)
The application of the partial nitritation and anammox process in municipal wastewater treatment
can convert them from energy consuming into energy producing process. Compared to conventional
biological nitrogen removal processes, the application of the partial nitriation and anammox process
can reduce the operation expenses by 60%, eliminates the need for external carbon sources and the
waste activated sludge is much lower [5]. Furthermore, the process reduces the greenhouse gas emissions by 90% since CO2 is consumed and there are no N2 O emissions [6]. Hydraulic retention time is
one of influencing factors for the anammox process [7, 8]. A practical purpose while applying anammox is to pursue a shorter HRT for higher nitrogen loading rate. So in this study, the authors used
PN and AX reactor to evaluate the effect of nitrogen treatment on domestic wastewater to meet the
requirements of the receiving source. The main purpose of the study was (i) to evaluate the effect of
nitrogen treatment on domestic effluent of the model system, (ii) to determine the appropriate water
retention time of the system.
2. Material and method
2.1. Partial nitritation (PN) and Anammoxreactor (AX) system
The PN + AX reactor system consists of Partial nitritation (PN) reactor and Anammox (AX)
reactor as shown in Fig. 1. The PN reactor [9] is rectangular in the bottom size of 10 × 20 (cm), height
31 cm, total volume V = 6.2L. Inside the PN reactor, there is a Felibendy material plate (16 cm ×
22 cm) implanted with Nitrosomonas bacteria contributed by Institute of Tropical Biology, Vietnam.
The AX reactor is a circular cylinder with an inner diameter of 7.1 cm, a height of 41 cm, a
useful volume of 1.62 liters [10, 11]. Within the reaction column using 1 × 1 × 0.8 cm Felibendy
cubes, anammox bacteria were cultured by the Meidensa company (Japan), using the Anammox strain
Candidatus Brocadia anammoxidans.
2.2. Wastewater and operating parameters
The study used the domestic wastewater from the three-compartment septic tank at the National
University of Civil Engineering. In order to simulate wastewater from the combined sewerage and
128

of 41 cm, a useful volume of 1.62 liters [10, 11]. Within the reaction column using
1×1×0.8cm Felibendy cubes, anammox bacteria were cultured by the Meidensa
company (Japan), using the Anammox strain Candidatus Brocadia anammoxidans.
Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

Thermostat

Felibendy
plates

Thermostat

119

valve

Inf.
tank

Eff.
PN/
Inf.
AX

Inf. Pump
200

Air

blower

94

Eff.
tank

80

300

Felibendy
cubes

Inf. Pump

73

AX reactor

PN reactor

Figure 1. Schematic diagram of Partial Nitritation and Anammox reactor system

Figure 1. Schematic diagram of Partial Nitritation and Anammox reactor system
drainage system in the rainy season and dry season and the separated sewerage system, wastewater
(Fontinchữ
trong1 hình
Times 1)
New

was diluted with gray water
the ratio
: 3 (period
and Roman)
1 : 2 (period 2). The non-diluted
wastewater
was
used
in
period
3
to
simulate
separated
sewerage
system.
Partial nitritation (PN) reactor
2. 2. Wastewater and operating parameters
was operated under aerobic conditions (DO ≈ 2 mg/l) and Anammox (AX) reactor under anaerobic
The study used the domestic wastewater from the three-compartment septic tank
conditions (DO < 0.5 mg/l).
at theThe
National
University
of Civil
Engineering.
to simulate
wastewater
from
PN reactor

is responsible
for the
conversion of In
partorder
of ammonium
to nitrite
to produce
nisuitable ratio
the Anammox
process.
orderseason
to take place
the partial
thetrite/ammonium
combined sewerage
andfor
drainage
system
in theInrainy
and dry
seasonnitritation
and the
by
Nitrosomonas
bacteria,
the
HRT
should
not
be

prolonged
due
to
that
ammonium
will
be
able1:3
to
separated sewerage system, wastewater was diluted with gray water in the ratio
transform to nitrate, but also should not be too short because of insufficient time for transformation
(period
and 1:2the
(period
2). conduct
The non-diluted
used
period
3 to
process.1)Therefore,
study will
experiments wastewater
with the HRTwas
in the
first in
period
(start-up
simulate
sewerage
Partial

nitritation
(PN) ofreactor
operated
period) isseparated
18h, then will
graduallysystem.
decrease to
12h and
9h. Composition
nitrogenwas
compounds
in
wastewater
andconditions
operating parameters
the PNand
+ AXAnammox
reactors system
is shown
in Table
under
aerobic
(DO ≈ 2ofmg/l)
(AX)
reactor
under1.anaerobic

conditions (DO<0.5 mg/l).
Table 1. Operating parameters of PN + AX reactor system
Inf. NH+4 – N

3Inf. NO2 – N

Inf. NO−3 – N

(mg/l)

(mg/l)

(mg/l)

PN

AX
12
9

−

HRT (h)

Period

Day to day

1a
1b

0-30
31-60

39.67±1.72

3.69 ± 0.29

1.18±0.53

18
12

2a
2b
2c

61-90
91-120
121-150

81.03 ± 1.38

4.95 ± 0.58

2.99±0.69

12
9
9

3a
3b

151-180
181-210

115.06±1.74

7.3±0.56

3.91±0.53

9
9

129

9
6
4.5
6
6

Com

Com


Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

2.3. Chemical analyses
The experiment was conducted in the laboratory of Water Supply and Sanitation Division, Faculty

of Environmental Engineering, National University of Civil Engineering. Parameters of influent and
effluent flow were measured 3 times per week. Ammonium concentrations were measured by colorimetic method with Nessler reagent at wavelength of 420 nm. In accordance with Standard Methods [12], nitrite and nitrate concentrations were estimated by the colorimetric method (4500-NO2 – B)
and the UV spectrophotometric screening method (4500-NO3 – B), respectively. Nitrite was known to
have an interfering response in the nitrate UV screening method of 25% of the nitrate response on a
nitrogen weight basis, thus the results were corrected by calculation. Levels of pH were measured by
using a Mettler Toledo-320 pH meter and DO was measured by using a DO meter (D-55, Horiba).
3. Result and discussion
3.1. Changes of ammonium (NH+4 – N), nitrite (NO−2 – N) and total nitrogen (TN) concentrations in
the partial nitritation reactor
As shown in Fig. 2, the first period was operated with diluted wastewater with an ammonium
concentration of 39.67±1.72 mg/l. In the first days of operation, Nitrosomonas bacteria was
not adapted to operating conditions, while competing with other microorganisms in domestic wastewater, the efficiency of NH+4 – N conversion to NO−2 – N is low. Ammonium concentration in wastewater
after the first 3 days of partial nitritation was only reduced from 38 mg/l to 26.25 mg/l, reaching a
conversion rate of 30.92%. However, in the following days, when the bacteria adhered, adapted and
promoted the role of converting ammonium to nitrite, the efficiency was significantly improved to
reach 51.79%. In addition, nitrite concentration formed in PN reactor was also increased, respectively,
from 12.07 mg/l (after the first day) to 19.62 mg/l (after day 30). As a result, the ratio of NO−2 – N:
NH+4 – N was also increased from 0.46 to 0.98. HRT in period 1b is reduced from 18h to 12h and
substrate concentration was kept as stage 1a. Results showed that the average conversion efficiency of
ammonium to nitrite was 51.48 ± 0.75% and after partial nitritation, the ratio of NO−2 – N: NH+4 – N
averaged 0.97 ± 0.05.
In the second period, the concentration of ammonium was increased from 39.67 ± 1.72 mg/l
to 81.03 ± 1.38 mg/l but the HRT was 12 hours as the first stage. Because of the increasing in
substrate concentration, the efficiency of the process was slightly reduced from 51.7% to 50.83%,
then stabilized toward the end of period 2a reaching 51.26%. The ammonium concentration after
the PN reactor was 39.66 ± 1.17 mg/l, the ammonium conversion efficiency of the partial nitritation
process during this period was 51.54 ± 0.71%. The ratio of NO−2 – N: NH+4 – N in wastewater after the
PN reactor was 1.02 ± 0.03. In the next 30 days of 2b and 2c period, the experiment was continued
running with the same substrate concentration but the HRT was reduced from 12h to 9h. With a HRT
of 9h, the ammonium removal efficiency of the PN model was 51.25 ± 1.13%, corresponding to the

ammonium concentration of 39.43 ± 1.12 mg/l in period 2a. Similar in period 2b, the ammonium
concentration of the outlet was 39.1 ± 0.45 mg/l, the average ammonium removal efficiency was
51.24 ± 0.71%. As a result, the efficiency of ammonium to nitrite conversion has decreased but not
significantly, so it can be confirmed that the 9h of HRT is appropriate for Nitrosomonas bacteria to
perform partial nitritation.
Thus, in the third period, the wastewater was collected after the septic tank (not diluted with
gray water) was used but experiments will conduct with HRT of 9h. The influent of the ammonium
NH+4 – N

130


also increased, respectively, from 12.07 mg/l (after the first day) to 19.62 mg/l (after
day 30). As a result, the ratio of NO2--N: NH4+-N was also increased from 0.46 to 0.98.
HRT in period 1b is reduced from 18h to 12h and substrate concentration was kept as
stage 1a. Results
showed
that
average
conversion
of ammonium to
Hanh, N. T.
M., Hoa, T.
T. H.the
/ Journal
of Science
and Technologyefficiency
in Civil Engineering
nitrite was 51.48 ± 0.75% and after partial nitritation, the ratio of NO2--N:
NH4+-N

concentration was 115.06 ± 1.74 mg/l and the effluent was collected at 56.51 ± 0.46
mg/l. Correaveragedto0.97
0.05. input and output are respectively 7.3 ± 0.56 mg/l and 58.55 ± 1.44 mg/l.
sponding
it, the± nitrite
The nitrite/ammonium ratio in wastewater after PN was 1.03 ± 0.02.
Changes of NH4-N in PN model
Period 1a
120 HRT=18h

Period 1b
HRT=12h

Period 2a
HRT=12h

Period 2b
HRT=9h

Period 2c
HRT=9h

Period 3b
HRT=9h

Period 3a
HRT=9h

60

110
50

90
80

40

70
60

30

50
40

20

30
20

Removal efficency(%)

Concentration (mg/l)

100

10

10

0

3
9
15
21
27
33
39
45
51
57
63
69
75
81
87
93
99
105
111
117
123
129
135
141
147
153
159
165

171
177
183
189
195
201
207

0

Time (days)
NH4-N influent

NH4-N effluent

NH4-N Removal efficency

(a) Changes of NH4 – N in PN model

(a) Changes of NH4-N in PN model
Changes of NO2-N in PN model

60

Period 1a
HRT=18h

Period 1b
HRT=12h

Period 2a
HRT=12h

Period 2b
HRT=9h

Period 2c
HRT=9h

Period 3a
HRT=9h

Period 3b
HRT=9h
100
90
80
70

40

60
30

50
40

20

30

Producing efficency(%)

Concentration (mg/l)

50

20

10

10
0
3
9
15
21
27
33
39
45
51
57
63
69
75
81
87
93
99

105
111
117
123
129
135
141
147
153
159
165
171
177
183
189
195
201
207

0

Time (days)
NO2-N influent

NO2-N effluent

NO2-N producing efficency

5
(b) Changes of NO2 – N in PN model

(b) Changes of NO2-N in PN model (Font chữ trong hình Times New Roman, không
đậm and nitrite in PN reactor
Figure 2. Changes of ammonium

Figure 2. Changes of ammonium and nitrite in PN reactor
In the second period, the concentration of ammonium was increased from 39.67
131
± 1.72 mg/l to 81.03 ± 1.38 mg/l but the HRT was 12 hours as the first stage. Because
of the increasing in substrate concentration, the efficiency of the process was slightly
reduced from 51.7% to 50.83%, then stabilized toward the end of period 2a reaching


ammonium levels are not high, ammonium removal efficiency is low, reaching only
49.33% due to nitrite to ammonium ratio has not yet met the ammonium oxidation
requirements of the Anammox process.

Commented [A5]: nitrite to ammonium ratio?

During the first 9 days of period 1a, NH4+-N ammonium concentration in effluent
was still
higher
10 mg/l.
in oftheScience
following
days, the ratio
of NO
2 -N:
Hanh,
N. T. than
M., Hoa,

T. T. However,
H. / Journal
and Technology
in Civil
Engineering
+
NH4 -N in the effluent of the anammox process was improved gradually. Ammonium
−
3.2. Changes
of ammonium
(NH+4 to
– N),
– N)remained
and total
nitrogen
concentrations
were reduced
less nitrite
than 10 (NO
mg/l2and
stable
at 8.78 ±(TN)
0.5 concentrations in
mg/l.
AX reactor
Simultaneously with the changes in ammonia concentration of the anammox
The effluent
from the PNwas
reactor
influent

flow
for the
AX reactor. In period 1a,
process,wastewater
the nitrite concentration
also was
reduced.
Nitrite
removal
efficiency
the partial increased
nitritation
process
was
effective
in
the
early
days
of
operation,
so
that wastewater from the
from 41.43% to 58.46% after 30 days of experiment. Total nitrogen removal
+
PN reactorefficiency
had an NH
–
N
concentration

of
26.25
mg/l
and
nitrite
concentration
is 12.07 mg/l. The
increased
from 38.38% to 52.69%, respectively.
4

+
effluent of the During
AX reactor
– Nreduction
concentration
of 13.3
as shown in Fig. 3. The
the nexthas
30 an
daysinitial
(periodNH
1b),4 the
of the HRT
in the mg/l
AX reactor
explanationfrom
for12h
thistois9h.
that

ammonium
levels removal
are not rate
high,
removal efficiency is
Byalthough
reducing the
HRT, the nitrogen
wasammonium
increased from
3
3
0.13
gN/m
.d
to
0.18
gN/m
.d,
ammonium
removal
efficiency
and
averaged
total
low, reaching only 49.33% due to nitrite to ammonium ratio has not yet met the ammonium oxidation
Commented [A6]: Please consider correcting
nitrogen
removal
efficiency

were 55.98 ± 3.72% and 51.33±1.4%, respectively.
requirements
of the
Anammox
process.
… and … respectively.”

Period 1a
HRT=12h

Concentration (mg/l)

25

Period 1b
HRT=9h

20
15
10
5
0

100
90
80
70
60
50
40

30
20
10
0

NH4-N influent

Removal efficiency (%)

30

NH4-N effluent
NO2-N influent
NO2-N effluent
NO3-N influent
NH4-N removal efficency
NO2-N removal efficency

3 6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60

45

100
90
80
100
9070
8060
7050
6040

5030
4020
3010
200

Removal efficiency (%)
Removal efficiency (%)

Changes
in Period
a. (a)
Changes
in Period
1 1

40

Concentration (mg/l)
Concentration (mg/l)

4535
4030
3525
3020
2515
2010
15 5
10 0
5
0

Period 2a
HRT=9h

Period 2a
HRT=9h

Period 2b
HRT=6h

7
Period 2c
HRT=4,5h

Period 2b
HRT=6h

Period 2c
HRT=4,5h

63 69 75 81 87 93 99 105 111 117 123 129 135 141 147

10
0

b. Changes in Period 2

NH4-N influent
NH4-N effluent

NH4-N influent
NO2-N influent
NH4-N effluent
NO2-N effluent
NO2-N influent
NO3-N effluent
NO2-N effluent
NH4-N removal
efficency
NO3-N effluent
NO2-N removal
efficency
NH4-N removal
efficency
NO2-N removal
efficency

63 69 75 81 87 93 99 105 111 117 123 129 135 141 147

60

100

55

90

50

Concentration (mg/l)

Concentration (mg/l)

60 45
55 40
50 35
45 30
40 25
35 20
30 15
25 10
20 5
15 0
10
5
0

80
100
70
90
Period 3a
Period 3b
60
80
HRT=6h
HRT=6h
50
70
40
Period 3a

Period 3b
60
30
HRT=6h
HRT=6h
50
20
40
10
Time (days)
30
0
153156159162165168171174177180183186189192195198201204207210 20

Time (days)

(c) Changes in Period 3

c. Changes in Period 3

153156159162165168171174177180183186189192195198201204207210

10
0

Removal efficiency (%)
Removal efficiency (%)

(b)Changes
ChangesininPeriod

Period 22
b.

NH4-N influent
NH4-N effluent

NH4-N
NO2-Ninfluent
influent
NH4-N
NO2-Neffluent
effluent
NO2-N
NO3-Ninfluent
effluent
NO2-N
NH4-Neffluent
removal
efficiency
NO3-N
NO2-Neffluent
removal
efficiency
NH4-N removal
efficiency
NO2-N removal
efficiency

FigureFigure
3. Changes

of nitrogen-containing
compounds
in AX
reactor
in each
period
3. Changes
of nitrogen-containing
compounds
in AX
reactor
in each
c. Changes
in Period 3
period

132

3. Changes
nitrogen-containing
compounds
reactor
each
InFigure
the second
period,ofammonium
concentration
into thein
PNAX
reactor

hasinincreased
period into the AX reactor has an ammonium
to 81.03 ± 1.38 mg/l and hence wastewater
concentration
39.43
± 1.12
mg/l and the
author conducted
3 periods
In the second
period,
ammonium
concentration
into theexperiments
PN reactor with
has increased
of
9h,
6h
and
4.5h.
The
results
showed
that,
with
the
HRT
of
4.5h,

the
ammonium
to 81.03 ± 1.38 mg/l and hence wastewater into the AX reactor has an ammonium


Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

removal/product rate (kgN/m3/d)

During the first 9 days of period 1a, NH+4 – N ammonium concentration in effluent was still higher
than 10 mg/l. However, in the following days, the ratio of NO−2 – N: NH+4 – N in the effluent of the
anammox process was improved gradually. Ammonium concentrations were reduced to less than 10
mg/l and remained stable at 8.78 ± 0.5 mg/l.
Simultaneously with the changes in ammonia concentration of the anammox process, the nitrite
concentration was also reduced. Nitrite removal efficiency increased from 41.43% to 58.46% after 30
days of experiment. Total nitrogen removal efficiency increased from 38.38% to 52.69%, respectively.
During the next 30 days (period 1b), the reduction of the HRT in the AX reactor from 12h to 9h.
By reducing the HRT, the nitrogen removal rate was increased from 0.13 gN/m3 .d to 0.18 gN/m3 .d,
ammonium removal efficiency and averaged total nitrogen removal efficiency were 55.98 ± 3.72%
and 51.33 ± 1.4%, respectively.
In the second period, ammonium concentration into the PN reactor has increased to 81.03 ± 1.38
mg/l and hence wastewater into the AX reactor has an ammonium concentration 39.43 ± 1.12 mg/l
and the author conducted experiments with 3 periods of 9h, 6h and 4.5h. The results showed that,
with the HRT
of 4.5h,
after taken
the AXfrom
reactor
16.45tank
± 1.5

mg/lthe
higher
In period
3, the
realammonium
domestic concentration
wastewater was
thewas
septic
with
than
the
allowable
level
of
the
receiving
source.
Meanwhile,
with
9h
and
6h
of
HRT,
the
ammonium
properties as shown in Table 1. After the partial nitritation process, wastewater with
concentrations were 9.45 ± 0.4 mg/l and 8.6 ± 0.55 mg/l, respectively. It can be seen that the time
average ammonium, nitrite, nitrate concentrations were 56.51±0.46 mg/l, 58.55±1.44

of 4.5h is too short for Anammox bacteria to process the metabolism. Therefore, it is necessary to
mg/l and 6.37±0.69 mg/l. With increasing inlet concentration, after 30 days, the
increase the HRT to 6h in the next experiment.
effluent
concentration
higher
thanseptic
the standard
variation
In periodammonium
3, real domestic
wastewaterwas
wasstill
taken
from the
tank with allowed,
the properties
as shown
from 10.89
mg/l
10.08nitritation
mg/l dueprocess,
to the recovering
theaverage
Anammox
bacterianitrite,
activity.
in Table
1. After
the to

partial
wastewaterof
with
ammonium,
nitrate
Besides, the
concentration
themg/l
effluent
is ±also
one With
of the
factorsinlet
concentrations
weresubstrate
56.51 ± 0.46
mg/l, 58.55 ±in1.44
and 6.37
0.69 mg/l.
increasing
concentration,
30 days,
the effluent
ammonium
still higher
than the
influencingafter
on the
treatment
efficiency

of the concentration
model. Afterwas
30 days
of period
3b, standard
the
allowed,
variation
from
10.89
mg/l
to
10.08
mg/l
due
to
the
recovering
of
the
Anammox
bacteria
effluent ammonium concentration was reduced to below 10 mg/l and remained stable
activity.
substrate
in the effluent
is also one
of the
factors influencing
on the

at theBesides,
range ofthe9.96
± 0.14concentration
mg/l. The effluent
total nitrogen
also
fluctuates
in the range
treatment efficiency of the model. After 30 days of period 3b, the effluent ammonium concentration
of 38.51 ± 0.91 mg/l.
was reduced to below 10 mg/l and remained stable at the range of 9.96 ± 0.14 mg/l. The effluent total
nitrogen The
also fluctuates
in thebetween
range of 38.51
0.91 mg/l.rate, nitrite removal rate, nitrate
relationship
TN ±removal
The relationship between TN removal rate, nitrite removal rate, nitrate production rate versus
production rate versus ammonium removal rate were established and shown in Fig.4.
ammonium removal rate were established and shown in Fig. 4.
0.60
0.50

TN removal rate
NO2 removal rate
NO3 product rate

y = 1.7644x
R² = 0.9953

0.40
0.30
y = 0.9861x
R² = 0.9915

0.20

y = 0.2217x
R² = 0.9212

0.10
0.00
0.03

0.08

0.13
0.18
0.23
NH4+-N removal rate (KgN/m3/d)

0.28

0.33

Figure
4. Ratios
of T-N removal,
NO−2 – N removal

andremoval
NO−3 – Nand
production
rates
to NH+4 – Nrates
removal
Figure
4. Ratios
of T-N removal,
NO2--N
NO3--N
production
to rates

NH4+-N removal rates
133

Ratios of T-N removal, NO2-N removal and NO3-N production rates to NH4-N
removal rates for Anammox reactor were 1.76:0.98:0.22, which are very similar to the
theoretical reaction ratios for the anammox reaction (Eq.2). In Hoa’s research [11],

Commente
it more clear.


Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

Ratios of T-N removal, NO2 – N removal and NO3 – N production rates to NH4 – N removal rates
for Anammox reactor were 1.76 : 0.98 : 0.22, which are very similar to the theoretical reaction ratios
for the anammox reaction (Eq. (2)). In Hoa’s research [11], ratios of T-N removal, NO2 – N removal

and NO3 – N production rates to NH4 – N removal rates for reactor 1 (MC 3-5mm diameter pieces)
were 1.98 : 1.15 : 0.17, for reactor 2 (MC 10-15mm diameter pieces) were 2.03 : 1.2 : 0.17, The
differences between theoretical ratio and the actual ratios for NO3 – N production may be due to the
biological denitrification process occur by existing of the anoxic heterotrophic bacteria which nitrate
was(combined
reduced to nitrogen
sewagegas.
and drainage system in rainy season, combined sewage and

drainage system in dry season, separated sewage and drainage system). The nitrogen
treatment efficiency of the system depends on the retention time of each reaction
The study
wasacarried
on PN
AXPN
reactor
system
3 periods corresponding
different
model.
With
HRT out
of 12h
in+the
reactor,
thewith
nitrite/ammonium
contenttoof
the
wastewater types of three sewage and drainage systems (combined sewage and drainage system in

effluent will not be suitable for Nitrosomonas bacteria in the PN reactor, hence the
rainy season, combined sewage and drainage system in dry season, separated sewage and drainage
treatment
efficiency
is veryefficiency
low, only
± 7,16%.
Aton
thethesame
time,time
the of
storage
system).
The nitrogen
treatment
of 45.54
the system
depends
retention
each retimemodel.
of 4.5h
in the
AX of
reactor
considered
to be
for Anammox
to
action
With

a HRT
12h inisthe
PN reactor,
the insufficient
nitrite/ammonium
content ofprocess
the effluent
so the
nitrogen treatment
of thehence
system
low at only
52.76 is
willtake
not place,
be suitable
fortotal
Nitrosomonas
bacteria inefficiency
the PN reactor,
theistreatment
efficiency
very± low,
onlyFor
45.54
7,16% time
as shown
in Fig. 5.
At thesystem
same time,

of 4.5h
in the
1.29%.
the±storage
in PN+AX
reactor
of 9 the
andstorage
6h, thetime
effluent
water
AXquality
reactor is
is considered
to
be
insufficient
for
Anammox
process
to
take
place,
so
the
total
nitrogen
ensured according to the requirements of the column B (none water supply
treatment efficiency of the system is low at only 52.76 ± 1.29%. For the storage time in PN + AX
purpose) according to QCVN 14:2008/BTNMT [1].

3.3. The efficiency of nitrogen treatment in PN + AX reactor system

130
120
110
100
90
80
70
60
50
40
30
20
10
0

Period 1a

Period 1b

Period 2a

Period 2b

Period 2c

Period 3a

Period 3b

100
90
80
70
60
50
40
30
20

Removal efficiency (%)

Concentration (mg/l)

reactor system of 9 and 6h, the effluent water quality is ensured according to the requirements of the
column B (none water supply purpose) according to QCVN 14:2008/BTNMT [1].

10
0
0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200 210
Time (days)
NO3-N effluent
NO2-N influent

NH4-N influent
NO2-N effluent

NH4-N effluent
'TN removal efficiency

Figure 5. Changes of nitrogen compounds in PN + AX reactor system

Figure 5. Changes of nitrogen compounds in PN+AXreactor system
result
ofgene
the gene
sequence
of bacteria
on carrier
materials
3.4.3.4.
TheThe
result
of the
sequence
of bacteria
on carrier
materials

Molecular biology techniques for bacteria identifying are carried out at the

Molecular biology techniques for bacteria identifying are carried out at the Center for BiotechnolCenter
for Biotechnology Research and Development, Institute of Biotechnology and
ogy Research and Development, Institute of Biotechnology and Food Technology, Hanoi University

Food Technology, Hanoi University of Science and Technology. The sequence
134 using Polymerase chain reaction (PCR);
includes designing bacteria-specific primers
Collected nucleotide sequence data were included in the Multalin comparison tool and

used Fast PCR software to reconcile the 16S rDNA gene segment on the mold
separated from the orginal sample (root) and from biomass material after use for real


Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

of Science and Technology and shown in Fig. 6. The sequence includes designing bacteria-specific
primers using
Polymerase
chain data
reaction
(PCR);
Collected
nucleotide
sequence
data were
obtained
nucleotide
sequence
were
included
in the
Multalin
comparison
toolincluded
to
in
the
Multalin
comparison

tool
and
used
Fast
PCR
software
to
reconcile
the
16S
rDNA
gene
identify conservative regions. From the results, about 30 nucleotides of head 5 'and 3' segment on the mold separated from the original sample (root) and from biomass material after use for
of the gene were used to carry out the design of the forward primer and the
real wastewater treatment. To conduct bait design, 10 16S rDNA sequences of the strain Candidacorresponding
reverse primer.
The selected
sequence
is included
in nucleotide
the FastPCR
tus Brocadia anammoxidans
were collected
from NCBI
data bank.
The obtained
sequence
software
to
calculate

the
parameters
and
select
the
sequence
that
satisfies
data were included in the Multalin comparison tool to identify conservative regions. From thethe
results,
about 30 nucleotides
head20-25
5’ and nucleotide
3’ of the genesequences,
were used tono
carryadditional
out the design
of the forward
requirements:
there ofare
pairing,
no
primer andpairing.
the corresponding
reverse primer.
The the
selected
sequence attached
is includedtointhe
theprimer

FastPCRissoftadditional
Primer sequence
together,
temperature
ware to calculate
the
parameters
and
select
the
sequence
that
satisfies
the
requirements:
there are
about 55-62oC.
20-25 nucleotide sequences, no additional pairing, no additional pairing. Primer sequence together,
the temperature attached to the primer is about 55-62◦C.
Afterobtaining
obtaining
therDNA-specific
16S rDNA-specific
After
the 16S
primer
1
2
3
4 M kb

pair
of
Candidatus
Brocadia
anammoxidans,
PCR
primer
pair
of
Candidatus
Brocadia
was
performed
to
amplify
the
16S
rDNA
gene
seganammoxidans, PCR was performed to amplify
– 3,0
ment using DNA mold separated from the previthe 16S rDNA gene segment using DNA mold
ous sample (root) and after use (from carrier maseparated
from
theareprevious
sample
– 2,0
terial). The
results
shown in Fig.

6. The(root)
resultsand
after
use (from
carrier
material).
results
obtained
DNA band
about
500 bp The
in both
sam- are
– 1,5
ples before
Theobtained
size obtained
was
shown
in Fig.and6. after
The use.
results
DNA
band
– 0,5
consistent
withintheboth
theoretical
sizebefore
according

about
500 bp
samples
and toafter
the design of the 16S rDNA gene fragment of the
use. The size obtained was consistent with the
strain Candidatus Brocadia anammoxidans. From
theoretical
to the design
of the
the results, size
it can according
be said that Candidatus
Brocadia
6. DNA electrophoresis
16S
rDNA gene
fragment
of the
strain Figure Figure
anammoxidans
strain still
exists in carrier
materi6. DNA electrophoresis of PCR products
of
PCR
products
amplify
the
16S

als after useBrocadia
to treat actual
domestic wastewater.
segment
from
DNA
Candidatus
anammoxidans.
From the amplify the 16S rDNA gene
rDNA
gene
segment
from
DNA
obtained
from
the
original
sample
(running
lines
This also
proves
Anammox
bacteria areBrocadia
comresults,
it can
bethat
said
that Candidatus

obtained
from
the
original
sample
1,2)
and
samples
after
being
used
for
water
pletely suitable for the actual domestic wastewater
anammoxidans strain still exists in carrier (running
treatment
(running
linessamples
3,4); M path,
lines
1,2) and
after
environment and play a role in the treatment of nistandard
DNA
ladder
materials
after
use
to
treat

actual
domestic
being
used
for
water
treatment
trogen in urban domestic wastewater.

wastewater. This also proves that Anammox (running lines 3,4); M path, standard
bacteria
are completely suitable for the actual DNA ladder.
4. Conclusions
domestic wastewater environment and play a role
The study used the Partial nitritation and Anammox reactor system to evaluate the removal effiin the treatment of nitrogen in urban domestic
ciency depend on the hydraulic rention time of nitrogen in dormitory’s wastewater from the National
wastewater.
University of Civil Engineering during 210 days. The partial nitritation reactor using Felibendy plate
4. with
Conclusion
Nitrosomonas bacteria and Anammox reactor using Felibendy cubes with presence of strains

Candidatus Brocadia anammoxidans achieved the stable treatment efficiencies over the time. The reTheconcluded
study used
the short
Partial
nitritation
andinAnammox
reactor
system

to evaluate
search
that the
HRT
of 4.5 hours
the AX reactor
affected
the Anammox
process
optimal hydraulic
retention
times forrention
PN and time
AX reactors
are 9h in
anddormitory’s
6h, respectively.
thenegatively.
removalThe
efficiency
depend on
the hydraulic
of nitrogen

wastewaterfrom the National University of Civil Engineering during 210 days. The
Acknowledgments
partial
nitritation reactor using Felibendy plate with Nitrosomonas bacteria and
Anammox
reactor

cubes with
of on
strains
Candidatus
This research
wasusing
carriedFelibendy
out in the framework
of the presence
Project “Study
the influence
of organic
Brocadia
achieved
the process
stable treatment
over the time.
matter to anammoxidans
anaerobic ammonium
oxidation
for nitrogenefficiencies
removal in wastewater”.
ThisThe
research
research concluded that the short HRT of 4.5 hours in the AX reactor affected the
Anammox process negatively. The optimal hydraulic
retention times for PN and AX
135
reactors are 9h and 6h, respectively.

Hanh, N. T. M., Hoa, T. T. H. / Journal of Science and Technology in Civil Engineering

is funded by National University of Civil Engineering (NUCE) under grant number 200-2018/KHXDTĐ. The authors would also like to acknowledge the Meidensa company, Nagoya, Japan, who distributed seed slugde; Kuraray Company, Japan, who supported Felibendy biomass carrier; Center for
Biotechnology Research and Development, Institute of Biotechnology and Food Technology, Hanoi
University of Science and Technology, who supported for bacteria identifying by Molecular biology
techniques.
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136