Study on Enhancement of Single Stage Autotrophic Biological

Denitrification System by Ultrasonic Wave

Li Zhang, Lei Zhang, Qi Jiang and Diannan Huang

School of municipal and environmental engineering, Shenyang Jianzhu University, Shenyang, Liaoning Province, China

Keywords:

Ultrasonic Strengthening, Canon, Extracellular Polymer.

Abstract: The application of one-stage autotrophic nitrogen removal is hampered by its overlong start-up and instability

under disturbance. A lab-scale batch tests were used to explore the optimal ultrasonic strengthening

parameters, which were determined based on changes in the activity of the Canon system and EPS. The

optimal ultrasonic intensity was 0.3 W/cm

, and the activity of Canon system was 40.68 mg N(gVSS

-1

The optimal ultrasonic time was 4 min, and the activity of Canon system was 41.02 mg N(gVSS

-1

. Under

the best ultrasonic intensity and ultrasonic time, it can be maintained for 6 days.

1 INTRODUCTION

The single-stage autotrophic biological nitrogen

removal process is one of the new biological nitrogen

removal processes promoted in recent years. It has the

advantages of low oxygen consumption, no need to

add organic carbon source, small reactor footprint,

etc. In addition, this process can improve nitrogen

removal efficiency, reduce nitrogen removal cost,

and develop the theory and engineering application of

nitrogen removal (Tang, 2015).

In order to reduce the possibility of instability of

nitrogen removal system and stimulate the growth of

Anammox bacteria, low-frequency ultrasonic with

mild action conditions, wide application range and no

secondary pollution was adopted in this study for

enhancement (Zhang, 2017). Ultrasonic wave is a

mechanical wave with multiple parameters, and the

corresponding parameters of different biological

nitrogen removal systems are not the same.

Therefore, it is of certain significance to explore the

influence of ultrasonic parameters on the efficiency

of single-stage autotrophic biological nitrogen

removal system.

Therefore, in order to increase the nitrogen

removal efficiency of the single-stage autotrophic

biological nitrogen removal system, low-frequency

ultrasonic batch test was used in this part to

strengthen the single-stage autotrophic biological

nitrogen removal system to study the optimal

working parameters. Fixed ultrasonic time, gradient

change of ultrasonic intensity, the effect of ultrasonic

intensity on biological denitrification system

strengthening efficiency, extracellular polymer and

each component were studied.

2 TEST MATERIALS AND

METHODS

2.1 Test Device and Operation

A certain amount of Canon system sludge was

inoculated and placed into 100 mL serum bottles, and

artificially simulated wastewater was added to the

100 mL scale line. The initial biomass concentration

of batch experiments was 1g VSS/L. The ultrasonic

generator has a fixed ultrasonic frequency of 25KHz,

and the power is between 40 and 200W.

The entire serum vial is covered with a black film.

Each serum flask was cultured in a shaker at

(30±1) ℃. Samples were taken every 12 hours with a

long syringe needle and the concentrations of NH

N, NO

-N and NO

-N in the water samples were

measured. The Canon system activity was calculated

based on the change of total nitrogen concentration

over time. Take the average in triplicate as the result.

2.2 Test Sludge Inoculation

In the ultrasonic enhancement stage, the inoculated

sludge was Canon sludge that was started in the

Zhang, L., Zhang, L., Jiang, Q. and Huang, D.

Study on Enhancement of Single Stage Autotrophic Biological Denitriﬁcation System by Ultrasonic Wave.

DOI: 10.5220/0011374500003443

In Proceedings of the 4th International Conference on Biomedical Engineering and Bioinformatics (ICBEB 2022), pages 957-961

ISBN: 978-989-758-595-1

957

laboratory. When the Canon system was started and

stable, the NLR was 0.84 kg N/(m

·d), and the

nitrogen removal rate was 0.68 kg N/(m

·d).

2.3 Test Water Quality

To prepare the synthetic wastewater, Ammonia

nitrogen is added to the mineral medium in the form

of NH

CI. The composition of the synthetic

wastewater was reported in Jin (Jin, 2013) et al.

3 RESULTS AND DISCUSSION

3.1 Influence of Different Ultrasonic

Intensity on the Activity of Canon

System

Figure 1: Activity of Canon system under different

ultrasonic intensities.

Five different ultrasonic intensity values were

selected from 0.1-0.5W/cm

, and 0.1W/cm

was

increased each time, and the fixed ultrasonic time was

3min. As shown in Figure 1, the activity of the Canon

system increased by a gradient when the ultrasonic

intensity was in the range of 0.1-0.4W/cm2 compared

to 0W/cm

. The enhancement peak was reached at

0.3W/cm

, when the activity of blank group was

31.91 mg N(gVSS

-1

, and that of 0.3W/cm

group

was 40.68 mg N(gVSS

-1

, 27.48% higher than that

of blank group. Compared with the group without

ultrasound enhancement, it increased by 27.48%. The

activity of Canon at 0.4 W/cm

was 32.74 mg

N(gVSS

-1

, which was lower than that at 0.3

W/cm

, but still higher than that of the group without

ultrasound enhancement. When the ultrasound

intensity was 0.5 W/cm

, the Canon activity

continued to decline, which was 5.73% lower than

that of the group without strengthening, indicating

that excessive ultrasound intensity would cause

damage to microorganisms, so the ultrasonic intensity

at this time had certain damage to the treatment effect

of the Canon system flora. The exchange rate

between matrix and cell was accelerated, and the

growth and metabolism of bacteria were promoted.

3.2 Influence of Different Ultrasonic

Intensity on EPS in Canon System

EPS is a kind of polymer organic matter secreted by

microorganisms, which mainly contains proteins,

carbohydrates, nucleic acids, humus, etc (Zhou,

2020).

In the sludge system, the main components of

EPS are polysaccharides and proteins, so the contents

of polysaccharides and proteins are mainly analyzed

in the study (Wu, 2013). As shown in Figure 2,

compared with the blank group, EPS content in the

enhanced group was increased. This indicates that

under the strengthening effect of the sludge system, a

certain amount of polysaccharides and proteins are

released from the microbial biofilm to reduce the

damage caused by external changes. When the

optimal ultrasound intensity was 0.3W/cm

, the

polysaccharide and protein contents were 25.07mg/g

VSS and 85.31mg/g VSS, respectively, which were

22.71% and 20.94% higher than those of the blank

group. When the ultrasonic intensity was 0.5W/cm

the contents of polysaccharide, protein, and total EPS

were significantly increased, and the contents were

32.22mg/g VSS, 122.65mg/g VSS, and 154.87mg/g

VSS, respectively, which were 57.71%, 73.87%, and

70.24% higher than those of the blank group.

Although ultrasonic enhancement can promote the

sludge treatment system and increase the secretion

rate of EPS, under the adjacent intensity gradient,

according to the normal secretion rate, the increase of

EPS content is larger when the ultrasonic intensity is

0.5W/cm

. The reason may be that the high-intensity

ultrasound causes excessive damage to

microorganisms, the cell wall is destroyed, and the

permeability of the cell membrane is increased,

leading to the release of a large number of

polysaccharides and proteins in cells, thus causing a

sudden increase in EPS content.

ICBEB 2022 - The International Conference on Biomedical Engineering and Bioinformatics

958

Figure 2: EPS content and composition under different

ultrasonic intensities.

3.3 Influence of Different Ultrasonic

Time on the Activity of Canon

System

Ultrasonic intensity is an important parameter in the

ultrasonic enhancement treatment of biological

systems, and ultrasonic time is also one of the

selected parameters. Pulse ultrasonic time refers to

the interval between two ultrasonic times, usually

several seconds; Continuous ultrasound time refers to

the duration of an ultrasound, usually a few minutes

to a few hours. In actual bioaugmentation, pulsed

ultrasound is rarely used, and most of it is continuous

(Yan, 2016).

Figure 3: Activity of Canon system after different

ultrasonic time.

When investigating the influence of ultrasonic

time on the activity of Canon system, the optimal

fixed ultrasonic intensity was 0.3W/cm

, and the

ultrasonic time was set to 0 min, 2 min, 4 min, 6 min,

8 min, and 10 min, respectively. The effect was

shown in Figure 3. The maximum activity of Canon

system was 41.02 mg N(gVSS

-1

at 4 min of

ultrasonic time, which was 31.25% higher than that

of the untreated group. After 4 min, the activity of the

Canon system did not continue to increase but began

to decrease. At 10 min, the activity of the Canon

system was 3.16% lower than that of the unenhanced

group, indicating that prolonged ultrasound time did

not further improve the activity of the Canon system,

but caused negative feedback and ultimately

decreased activity. Schlafer (Schläfer, 2002) also

believed that stimulation of the biological system

within a certain range can improve microbial activity,

and when the range is exceeded, it will cause

inhibition.

3.4 Influence of Different Ultrasonic

Time on EPS in Canon System

Studies on the effects of ultrasound on other

microorganisms show that low-intensity ultrasound

can cause micro-damage to cells and destroy the

structure of the biological walls and cell membranes,

in which microorganisms produce more EPS to

protect themselves from adverse environmental

conditions. In order to explore the influence of

ultrasonic time on the activity of Canon system, the

contents of polysaccharide, protein, and total EPS at

the fixed ultrasonic intensity of 0.3W/cm

and

ultrasonic time of 0 min, 2 min, 4 min, and 10 min

were selected to study.

Figure 4: EPS content and composition after different

ultrasonic time.

The result is shown in Figure 4. When the short-

time ultrasound time was 2 min and 4 min, the total

EPS content increased gradually. When the short-

time ultrasound time was 4 min, the polysaccharide,

Study on Enhancement of Single Stage Autotrophic Biological Denitriﬁcation System by Ultrasonic Wave

959

protein, and total EPS contents were 26.64mg/g VSS,

87.91mg/g VSS, 114.55mg/g VSS, respectively.

Compared with the group without ultrasound

enhancement, it increased by 26.50%, 21.32%, and

22.49%, respectively. When the ultrasound time was

10min, the total EPS showed an obvious increase

process, and the contents of polysaccharide, protein,

and total EPS were 33.56mg/g VSS, 138.05mg/g

VSS and 171.61mg/g VSS, which increased by

59.35%, 90.52%, and 83.50% compared with the

group without ultrasound enhancement.

3.5 Activity of Canon System under

Different Ultrasonic Cycles

Prolonged ultrasound can exceed the tolerance limit

of microorganisms and cause irreversible cell

damage. In order to maximize the enhancement effect

of low intensity ultrasound, the ultrasonic cycle was

studied after the optimal ultrasonic intensity and

ultrasonic time were discussed. In this part of the

study, the ultrasonic intensity was set at 0.3W/cm

and the ultrasonic time was set at 2min and 6min

respectively. Based on the optimal ultrasonic time of

3min, the maximum number of active maintenance

cycles of the Canon system was studied.

Figure 5: Changes in the activity of the Canon system under

the optimal ultrasonic intensity and different ultrasonic

time.

Under the conditions of ultrasonic intensity of

0.3W/cm

and ultrasonic time of 0min, 2min, 4min

and 6min, the activity test results of each Canon

system are shown in Figure 5. The test period shall be

divided into 48 hours each. In the second cycle, the

activity of the test group was close to that of the blank

group when the ultrasonic time was 2min and 6min.

Therefore, it was proved that the effect maintained for

two cycles when the ultrasonic time was 4min and

6min. However, when the ultrasonic time was 4min,

the activity of the Canon system was similar to that

of the untreated group only in the third cycle. This

indicates that when the ultrasonic intensity is

0.3W/cm

and the ultrasonic time is 4min, the

maximum period can be maintained for 6days, which

is one more period than the 2min and 6min test

groups, indicating that the Canon system has certain

adaptability to ultrasonic radiation and can promote

the activity of the Canon system in a relatively short

period of time (Liu, 2003).

3.6 Influence of Different Ultrasonic

Periods on EPS in Canon System

Figure 6: Comparison of EPS content and composition

between ultrasound and non-ultrasound treated Canon

systems.

Figure 6 shows the comparison of polysaccharide,

protein, and total EPS contents between the

ultrasound enhanced group and the non-enhanced

group after the ultrasonic intensity of 0.3W/cm

and

ultrasonic time of 4 min. On the first day after

ultrasonic enhancement, the contents of

polysaccharide, protein and total EPS in the

experimental group were 63.39mg/g VSS,

158.48mg/g VSS, and 221.87mg/g VSS, respectively,

increased by 28.63%, 32.69%, and 31.53% compared

with those in the group without ultrasonic

enhancement, respectively. On the third day after the

completion of the enhancement, the growth rate of

each component decreased, but the contents of

polysaccharide, protein, and total EPS in the

experimental group were still higher than those in the

non-ultrasonic enhancement group. The production

rates of polysaccharide, protein, and total EPS were

13.39 mg/g VSS/d, 31.97 mg/g VSS/d and 45.36

mg/g VSS/d, respectively, 1-3 days after ultrasonic

ICBEB 2022 - The International Conference on Biomedical Engineering and Bioinformatics

960

treatment. On the sixth day of ultrasound treatment,

the contents of all components in the ultrasound

enhanced group and the group without ultrasound

were basically the same, but the production rates of

polysaccharide, protein, and total EPS decreased to

4.11 mg/g VSS/d, 9.51 mg/g VSS/d and 13.62 mg/g

VSS/d during 4-6 days of ultrasound treatment,

respectively. The results showed that low intensity

ultrasonic stimulation increased the metabolic rate of

cells, and more metabolites were excreted out of cells

(Pitt, 2003).

4 CONCLUSIONS

(1) In the range of 0.1-0.4 W/cm2, the activity of the

Canon system was higher than that of the blank

group, and the maximum value of 40.68 mg N(gVSS-

1) d-1was reached at 0.3W/cm2, which was 27.48%

higher than that of the blank group. When the

ultrasound intensity was 0.5W/cm2, the activity was

5.73% lower than that of blank enhancement group.

(2) In the ultrasonic time range of 2-8min, the

activity of the Canon system was higher than that of

the group without ultrasound enhancement, and the

maximum enhancement was 41.02 mg N(gVSS

-1

) d

-1

at the ultrasonic time of 4 min, which was 31.25%

higher than that of the group without ultrasound

enhancement. The activity of the Canon system was

3.16% lower than that of the untreated group.

(3) when the optimal ultrasonic intensity was

0.3W/cm

and the optimal ultrasonic time was 4 min,

the maintenance time of ultrasonic enhancement was

6 days, one cycle longer than that of adjacent

ultrasonic time of 2 min and 4 min. On day 1 after

ultrasonic treatment, the content of polysaccharide,

protein, and total EPS in the Canon system was

63.39mg/g VSS, 158.48 mg/g VSS, and 221.87 mg/g

VSS, respectively, which increased by 28.63%,

32.69%, and 31.53% compared with the group

without ultrasonic enhancement, respectively. With

the extension of time, the secretion rate of each

component decreased, and the ultrasonic enhanced

group was similar to the unenhanced group on day 6.

ACKNOWLEDGEMENT

This research was financially supported by General

project of scientific Research Plan of Education

Department of Liaoning Province (LJKZ0601)

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