Industrial Water Treatment | Performance Study of EGSB Reactor in Treating Wastewater from Antibiotic Pharmaceutical Manufacturing

Antibiotics are commonly used biological products in China, and their production generates a large amount of wastewater. This wastewater contains high concentrations of organic matter and is somewhat toxic. Improper disposal can cause significant harm to the surrounding environment.

Currently, anaerobic biological treatment is a common method for treating high-concentration organic wastewater. This method removes organic pollutants while also recovering some methane. The Expanded Granular Sludge Bed ( EGSB) reactor is a new anaerobic treatment technology. Compared with the commonly used Upflow Anaerobic Sludge Blanket ( UASB ) reactor, it has advantages such as high upflow velocity, high organic matter removal rate, strong shock resistance, and high volumetric loading rate. It can be widely used in the treatment of high-concentration or toxic and refractory organic wastewater.

This study constructs an EGSB reactor to treat high-concentration antibiotic pharmaceutical wastewater. The study investigates its treatment performance and shock resistance, analyzes the microbial community structure to identify dominant bacterial populations, and aims to provide technical support for future EGSB reactor applications in the treatment of antibiotic pharmaceutical wastewater.

01

Experimental Section

1
Experimental Wastewater

The substrate for this experiment was antibiotic pharmaceutical wastewater collected from the collection pool of a pharmaceutical factory's wastewater treatment plant. The collected wastewater was stored in plastic containers and used at room temperature. The wastewater quality indicators were tested as follows: COD was (15772±225) mg/L, SS was (478±32) mg/L, NH ₄⁺ -N was (17±2) mg/L, TP was (3.9±0.6) mg/L, alkalinity was (235±12) mg/L, pH was 6.8±0.3, and color was 752±56 times. This wastewater contains high concentrations of organic matter, and the nitrogen and phosphorus nutrients are insufficient,

therefore, a certain amount of NH ₄ Cl and KH ₂ PO ₄ were added to the influent to maintain the influent m(C):m(N):m(P) ratio of 500:5:1 to meet the basic metabolic needs of anaerobic microorganisms.

2
Inoculum Sludge

The reactor inoculum sludge was collected from a municipal wastewater treatment plant. It was dewatered sludge from a plate-and-frame filter press in the dewatering room (water content of 75%). This sludge mainly came from the effluent sludge of the secondary sedimentation tank after the biochemical reaction. After collection, the sludge was screened using a 0.600 mm × 0.450 mm (30 mesh × 40 mesh) stainless steel screen to remove large particles to prevent blockage of the peristaltic pump and pipes. The pretreated sludge was directly added to the EGSB reactor. The total suspended solids (TS) and volatile suspended solids (VS) at the time of inoculation were (9.6±0.4) g/L and (7.2±1.0) g/L, respectively.

3
Experimental Setup

The EGSB reactor setup is shown in Figure 1.

Figure 1 EGSB reactor setup

The EGSB reactor was made of organic glass with an effective volume of 25 L. A fiberglass gas-liquid-solid three-phase separator was installed at the top, and a pH probe was installed inside to monitor system pH changes in real time. Resistance wires were wrapped around the reactor wall for electric heating, and it was interlocked with a temperature probe to automatically control the reaction temperature at (35±1) ℃. After the pretreated inoculum sludge was

added to the EGSB reactor, the wastewater was transported into the reactor by a variable-speed peristaltic pump to dilute the influent COD, and the initial volumetric loading rate was controlled at 1.0 kgCOD/(m ³ ·d) (HRT was 48 h). During the start-up operation of the reactor, the volumetric loading rate was gradually increased until full-load operation [7.9 kgCOD/(m ³ ·d)]. After the reactor was successfully started up, the system HRT was sequentially reduced to 40 and 32 h [corresponding volumetric loading rates were 9.5 and 11.8 kgCOD/(m ³ ·d), respectively] to determine the optimal volumetric loading rate. Part of the reactor effluent was recirculated to the influent via a metering pump.

4
Detection Methods

The volume of biogas produced by the reactor was measured using an LMH-1 wet gas flow meter (Shandong Sangze Instrument). The methane volume fraction was measured using a 7890B gas chromatograph (Agilent Technologies, USA).

COD, BOD5, TS, VS, NH ₄⁺ -N, TP, alkalinity, and color were analyzed using national standard methods. Sludge particle size was determined using the wet sieving method, and the analysis of bacterial community types was performed using the centrifugation washing, reagent extraction, gel electrophoresis, and gene amplification methods described in the literature.

02

Results and Discussion

1
EGSB Reactor Operating Performance

The continuous operating performance of the EGSB reactor was investigated over the entire experimental period ( 200 d), and the results are shown in Figure 2.

Figure 2 Experimental operating performance of the EGSB reactor

From Figure 2, it can be seen that during the start-up of the EGSB reactor, the volumetric loading rate was gradually increased from 1.0 kgCOD/(m ³ ·d) to 7.9 kgCOD/(m ³ ·d) (full load) by increasing the influent COD.

Under each volumetric loading rate condition, when When the COD removal rate reached 80%, the volumetric load was increased successively to 2.0, 2.9, 3.9, 4.9, 5.8, 6.9, and 7.9 kgCOD/(m ³ ·d). The entire startup process lasted about 90 days. After successful startup, the performance of the EGSB reactor was stable within 22 days of operation, with an average COD removal rate of 91.6% ± 1.8%, and an average effluent COD of (1320 ± 15) mg/L.

In addition, During the startup of the EGSB reactor, each time the volumetric load was increased, the average COD removal rate of the system showed a trend of first decreasing and then increasing. This is because the anaerobic microorganisms in the reactor need a certain adaptation period to the new operating conditions.

On the 102nd day of reactor operation, the system HRT was reduced to 40 h to increase the volumetric load to 9.5 kgCOD/(m ³ ·d). After 20 days of operation, it was found that the COD removal rate of the system had hardly changed, with an average COD removal rate of 91.4% ± 0.8%. When the system HRT was further reduced to 32 h [at this time, the volumetric load was 11.8 kgCOD/(m ³ ·d)], the average COD removal rate decreased to 83.5% ± 1.2%, and the average effluent COD increased to (2558 ± 97) mg/L.

At the same time, The presence of granular sludge was found in the EGSB effluent. This is because the excessively fast upflow velocity at HRT of 32 h led to the loss of some sludge, thus reducing the number of anaerobic microorganisms and decreasing the system treatment efficiency. The system HRT was increased again to 40 h, and after 20 days of operation, the system operating performance recovered to the original level, and the average COD removal rate increased to 91.7% ± 1.1%.

This shows that in this study, the EGSB reactor is more advantageous when operating at an HRT of 40 h [volumetric load 9.5 kgCOD/(m ³ ·d)], achieving both high COD removal capacity and reduced reactor volume load.

During the reactor operation from day 1 to 157, the pH in the EGSB reactor remained stable at 6.9–7.6, a pH range conducive to the proliferation and metabolism of anaerobic methanogenic bacteria. To investigate the stability and shock resistance of the reactor operation, on the 158th day of operation, a certain amount of 31% hydrochloric acid was added to the influent to reduce the pH to 5.6, below the optimal pH range (6.5–8.5) for anaerobic microorganisms, to investigate the operating effect of the EGSB reactor.

The results show that when the influent pH decreased, the average COD removal rate of the EGSB reactor immediately decreased significantly, resulting in an increase in effluent COD. The average COD removal rate decreased to 74.8% ± 1.9%, a decrease of 18.4% compared to before pH adjustment, and the effluent COD increased to (3973 ± 102) mg/L.

As time went on, the anaerobic microorganisms gradually adapted to the low pH environment, and the COD removal rate gradually increased and reached a stable state. After 25 days of continuous operation of the EGSB reactor, the average COD removal rate recovered to 83.4% ± 1.3%, lower than the average COD removal rate (91.7% ± 1.1%) before pH adjustment. This indicates that the low pH environment had a certain degree of irreversible negative impact on the methanogenic bacteria in the reactor.

Therefore, maintaining the system pH within the optimal range for anaerobic microorganisms is crucial for the efficient and stable operation of the EGSB reactor.

2
Methane production performance of the EGSB reactor

The changes in methane production rate and methane volume fraction during the operation of the EGSB reactor are shown in Figure 3.

Figure Figure 3. Changes in methane production of the EGSB reactor

From Figure As can be seen from Figure 3, during the startup of the EGSB, the methane production rate gradually increased with the gradual increase of the volumetric load. When the system was operating at full load, the average methane production rate was (2.33 ± 0.04) L/(L·d). When the system HRT was reduced to 40 h, the methane production rate increased to (2.56 ± 0.05) L/(L·d). As the HRT was further reduced to 32 h, the methane production rate continued to increase to (2.82 ± 0.04) L/(L·d).

Although at HRT of 32 h, the average COD removal rate of the system was lower, at 83.5% ± 1.2%, the total amount of organic matter in the influent was higher, and the total amount of organic matter removed was higher than that under the operating conditions of HRT of 48 h and 40 h. Therefore, the system produced more methane.

However, considering the COD removal efficiency, the system HRT of 32 h is not suitable for the long-term operation of the EGSB reactor. When the system was subjected to low pH shock, the COD removal rate decreased significantly, and the methane production rate also decreased significantly to (2.04 ± 0.04) L/(L·d), a decrease of 27.7% compared to before pH adjustment. The negative impact of low pH on methanogenic bacteria led to a decrease in methane production.

As the anaerobic microorganisms gradually adapted to the new metabolic environment, their metabolic performance gradually increased and stabilized, and the methane production rate gradually increased, eventually stabilizing at about (2.26 ± 0.07) L/(L·d), lower than the methane production rate before pH adjustment.

During the entire operation process, the methane volume fraction in the biogas produced by the EGSB reactor was relatively stable, varying between 57.3% and 72.2%. The methane volume fraction was less affected by the volumetric load and pH shock. In addition to methane, the biogas also contained a certain amount of CO ₂ and a small amount of H ₂ etc.

3
Formation of granular sludge in the EGSB reactor

To ensure the efficient and stable operation of the anaerobic reactor, the formation of granular sludge is crucial. Compared with flocculent sludge, granular sludge has better settling properties and microbial density, which can improve the system's resistance to external factors (such as load, Interference ability of pH and toxic elements, etc. In previous studies, granular sludge was defined as sludge with a particle size ≥0.5 mm. The changes in the proportion of sludge with different particle sizes during the startup process of the EGSB reactor were investigated, and the results are shown in Figure 4.

Figure 4 Changes in the proportion of sludge with different particle sizes during the startup of EGSB

From Figure As can be seen from Figure 4, the raw sludge is basically flocculent sludge, with only 18.3% of the sludge having a particle size ≥0.5 mm, very little granular sludge, and SV30 of (33.1±0.4), indicating poor settling performance. As the reactor starts to operate, the proportion of granular sludge gradually increases, and the diameter of the granular sludge gradually increases.

When the reactor operates to the 30th day, the system begins to show granular sludge with a particle size ≥2 mm, and the proportion of granular sludge increases from 13.7% of the raw sludge to 55.2%, indicating that granular sludge domestication is underway.

On the 60th day of operation, granular sludge continued to increase, with the proportion increasing to 75.7%, of which granular sludge with a particle size ≥1 mm and a particle size ≥2 mm accounted for 18.4% and 7.7%, respectively.

After the successful startup of the reactor (the 90th day), the proportion of granular sludge can reach 80.7%, which is 6 times that of the raw sludge. This is also the reason why the reactor has a high COD removal rate and methane yield under full-load operating conditions.

After the successful startup of the EGSB reactor, the proportions of sludge with particle sizes of 0.5~0.7 mm, 0.7~1.0 mm, 1.0~2.0 mm, and ≥2 mm were 36.5%, 21.4%, 12.6%, and 9.7%, respectively.

Meanwhile, the detection of granular sludge SV30 is 21.3±0.4, which is lower than that of raw sludge, indicating that granular sludge has higher settling performance. In addition, the VS of granular sludge is as high as (39.1±1.5) g/L, which is much higher than that of raw sludge (7.2±1.0) g/L, indicating that granular sludge has higher biomass.

4
Analysis of anaerobic microbial community structure

For Sampling and analysis of microbial community structure and relative abundance were conducted on the 90th and 200th days of operation of the EGSB reactor, and the results are shown in Figure 5.

Figure 5 Analysis of anaerobic microbial community structure and relative abundance

In the anaerobic biological treatment process, microorganisms are mainly divided into two categories: acid-producing bacteria and methanogenic bacteria. Acid-producing bacteria are mainly responsible for converting organic compounds in water into volatile organic acids and a small amount of alcohols through hydrolysis and acidification. In this study, The main volatile organic acids in the effluent of the EGSB reactor are acetic acid and butyric acid.

From Figure As can be seen from Figure 5, after the successful startup and stable operation of the EGSB (90th day), the main dominant acid-producing bacteria were Geobacter, with a relative abundance of 20.1%±2.2%. The main metabolic products of this bacteria are acetic acid and butyric acid.

Methanomassiliicoccus is the dominant methanogenic bacteria, playing a major role in the methanogenesis process, with a relative abundance of 31.2%±1.8%. Methanomassiliicoccus is one of the common bacteria in mesophilic anaerobic biological treatment processes, mainly converting volatile organic acids produced by acid-producing bacteria into methane and carbon dioxide, etc.

The dominant bacterial community structure in this study is consistent with The research results of C. M. Chen et al., who constructed a UASB to treat coal gasification wastewater and analyzed the anaerobic microbial community structure, found that the dominant bacteria were Geobacter and Methanomassiliicoccus, and the anaerobic biological treatment process was mainly completed by the above two bacteria. After the system was subjected to low pH impact and stable operation (200 days), there was no significant change in the structure of each microbial community, but the relative abundance changed to varying degrees.

Dominant methanogenic bacteria The relative abundance of Methanomassiliicoccus decreased from 31.2%±1.8% to 18.7%±1.6%, and this bacteria was significantly affected by low pH, which is the main reason for the decrease in COD removal rate of the system.

Conversely, The relative abundance of Methanothrix increased from 4.8%±0.2% on the 90th day to 10.5%±0.5%, indicating that this methanogenic bacteria has excellent tolerance to pH changes and plays an important role in stable operation under low pH conditions.

Previous studies have shown that Methanothrix can perform metabolism under lower pH conditions with good metabolic performance. In addition, the relative abundance of other methanogenic bacteria decreased to varying degrees.

After low pH impact, the relative abundance of the dominant acid-producing bacteria Geobacter increased from 20.1%±2.2% to 28.9%±1.1%, which is related to the metabolic environment of acid-producing bacteria suitable for low pH conditions. The metabolic products of acid-producing bacteria are mainly volatile organic acids, which will make the metabolic environment acidic, which determines its characteristic of tolerating low pH impact.

03

Conclusion

（ 1) The EGSB reactor has certain feasibility for treating high-concentration antibiotic pharmaceutical wastewater. Under the optimal volumetric load of 9.5 kgCOD/(m ³ ·d) (HRT 40 h), the average COD removal rate can reach 91.4%±0.8%, and the methane yield is (2.56±0.05) L/(L·d).

（ 2) Although pH changes have a significant impact on the stable operation of the EGSB reactor, it still has a certain ability to resist low pH impact. After being subjected to low pH impact, the COD removal rate of the EGSB reactor can still reach 83.4%±1.3% after a certain adaptation period.

（ 3) The dominant bacteria in the EGSB reactor are Geobacter and Methanomassiliicoccus. After being subjected to low pH impact, the dominant bacterial community structure did not change, but the relative abundance of Geobacter increased while the relative abundance of Methanomassiliicoccus decreased. This indicates that low pH has a certain negative impact on methanogenic bacteria.