Skip to main content
NCBI home page
Scientific Reports logoLink to Scientific Reports
. 2026 Apr 9;16:16196. doi: 10.1038/s41598-026-45194-w

Efficacy and limitations of the A2O process in simultaneous removal of surfactants and nutrients from municipal wastewater

Tahereh zarei Mahmoudabadi 1,2, Fahimeh Teimouri 2,✉, Amir Hossein Bagheri 1,2, Sara Jambarsang 3
PMCID: PMC13201829  PMID: 41957102

Abstract

This study evaluated the efficacy of the Anaerobic-Anoxic-Oxic (A2O) process for the simultaneous removal of organic pollutants, specifically surfactants (detergents), and nutrients (nitrogen and phosphorus) from municipal wastewater. Sampling was conducted over a 12-month period from September 23, 2023 to September 22, 2024, at a monthly frequency. During each sampling event, grab samples were collected from multiple points within the treatment train: the raw influent wastewater, the influent and effluent streams of the anaerobic, anoxic, and aerobic tanks, and the final effluent from the secondary clarifier. The anionic surfactants (as methylene blue active substances, MBAS), ammonium-nitrogen (NH₄⁺-N), nitrate-nitrogen (NO₃⁻-N), nitrite-nitrogen (NO₂⁻-N), and total phosphorus (TP)—were quantified in accordance with Standard Methods for the Examination of Water and Wastewater. The mean final effluent concentrations were 0.22 ± 0.05 mg/L for linear alkylbenzene sulfonate (LAS), 13.18 ± 3.7 mg/L for nitrate (as NO₃-N), and 4.46 ± 0.18 mg/L for phosphorus (as P). Process performance data indicated that the oxic tank was the primary site for LAS and phosphorus removal, while the anoxic tank facilitated the most significant nitrate reduction. While the Anaerobic-Anoxic-Oxic (A2O) process proficiently removes surfactants and phosphorus via microbial metabolism, its performance in nitrate elimination is constrained. Inadequate denitrification in the anoxic stage, frequently due to a limited biodegradable carbon source or improper redox conditions, indicates a need for process optimization.

Keywords: Wastewater treatment, Nutrients, Surfactants, Biological process

Subject terms: Chemistry, Environmental sciences, Water resources

Introduction

Human activities have led to significant increases in nitrogen and phosphorus in the biosphere, altering biological communities in aquatic ecosystems, damaging drinking water supplies, and threatening the growth of jobs and economic sectors that require high-quality and sustainable water resources1. In aquatic ecosystems, nitrogen and phosphorus act as fertilizers, leading to eutrophication, which increases the growth of harmful aquatic plants and harmful algal blooms, leading to the production of toxins that threaten human, animal, and livestock health. Human sources such as sewage discharge, fossil fuel combustion, agricultural and urban runoff, and landfill leachate have increased the input of nutrients (nitrogen and phosphorus) into the environment, causing eutrophication and imposing economic costs of nearly 2.2 billion dollars per year in the United States2,3.

The discharge of nitrogen compounds (N) such as organic nitrogen (urea, amines, and protein) and inorganic nitrogen (ammonia/ammonium, nitrite, and nitrate) increases nitrate concentrations in groundwater due to the high mobility of nitrate4. Nitrate is a precursor to forming N-Nitroso compounds (NOCs) through endogenous nitrosation. Most NOCs are carcinogenic and teratogenic5. Nitrate is readily absorbed in the upper gastrointestinal tract and distributed throughout the human body. Once it reaches the salivary glands, it is actively transported from the blood into the saliva, and its level may be up to 20 times higher than in plasma. According to the World Health Organization (WHO), the permissible limit of nitrate concentration in drinking water should not exceed 50 mg NO₃⁻/L (11 mg NO₃-N/L). This limit is set to protect against methemoglobinemia in infants, also known as blue baby syndrome, the most well-known health consequence of exposure to high nitrate levels6. Shaban et al. reported that, out of 78 infants, 25 developed methemoglobinemia associated with nitrate-contaminated water at concentrations ranging from 58.3 to 178.4 mg/L7.

Phosphorus exists in wastewater in both soluble and insoluble forms. In municipal wastewater, approximately 50 to 70% of the total phosphorus consists of orthophosphate, while the remaining 30 to 50% is made up of polyphosphates and organic phosphorus. Orthophosphate concentration of in municipal wastewater typically ranges from 4 to 16 mg/L8,9. According to the US Environmental Protection Agency, the global annual cost of controlling and reducing phosphorus pollution in freshwater resources is estimated at around 265 billion dollars annually10. Phosphate, when present in foods, is absorbed in the intestines, and excessive levels can cause chronic kidney disease and reduce the kidneys’ ability to excrete phosphate. This ultimately results in phosphate accumulation in the body, which acts as a catalyst for extensive damage to cellular components, bones, and cardiovascular structures11.

Detergents are a significant and influential pollutant in wastewater. Their surfactant properties lead to the formation of harmful foams in water sources. Surfactants are the primary ingredients used in the production of synthetic detergents and are categorized into four types: anionic, cationic, nonionic, and amphoteric12. Anionic surfactants are part of human life, widely used in homes and industries as an active ingredient in soaps and detergents13. The per capita consumption of synthetic detergents is increasing due to their cleansing and antimicrobial properties, which are applied in hygiene and infectious disease control11. In the 1930s, alkylbenzene sulfonates were first produced as branched alkylbenzene sulfonates (BAS). Subsequently, due to their poor biodegradability and environmental concerns, they were replaced in the 1960s by linear alkylbenzene sulfonates (LAS). Linear alkylbenzene sulfonates (LAS) constitute 40% of anionic surfactants14. Table 1 summarizes the concentrations of key detergent-derived surfactants reported in raw municipal wastewater.

Table 1.

Reported concentrations of surfactants in raw municipal wastewater.

Surfactant type Name Concentration References
Anionic surfactants Linear alkyl benzene sulfonate (LAS) 30 mg/L 14
1.8 mg/L 15
15.8 mg/L 16
10–20 mg/L 17
1–10 mg/L 18
Sodium dodecyl sulfate (SDS) 1.8 mg/L 19
Cationic surfactants Quaternary ammonium compounds (QACs) 0.5 mg/L 20
500 µg/L 21
Nonionic surfactants Nonylphenol ethoxylate (NPEO) 4.12 mg/L 22
Open in a new tab

As shown, anionic surfactants, particularly linear alkylbenzene sulfonates (LAS), are typically present at the highest concentrations compared to other surfactants.

The A2O process is an advanced biological treatment process with a three-stage system: anaerobic, anoxic, and oxic. In the anaerobic stage, microorganisms, especially phosphorus-removing bacteria, consume organic matter as a carbon source to obtain energy. In the anoxic stage, the nitrate produced in the oxic stage is placed in an oxygen-limited environment. In this stage, microorganisms use nitrate as an electron acceptor and convert it into nitrogen gas released into the atmosphere. In the oxic stage, oxygen is injected into the environment so aerobic microorganisms can decompose residual organic matter and ammonium23,24. As a result, the A2O process is a promising system for the simultaneous removal of nutrient and organic pollutants. The A2O process has high flexibility, low sludge production, and minimal odor emissions, making it one of the most efficient biological treatment options25,26. Numerous studies have been conducted on the removal of various pollutants from wastewater using the A2O process. Villanueva et al. achieved 98.5% removal efficiency for COD at a C/N ratio of 5 using the A2O process27. Zhou et al., in their investigation of the microbial performance of the A2O system for the treatment of electroplating wastewater, reported averages of 28, 0.27, and 10 mg/L in the effluent for COD, phosphorus and TN respectively28. The previous researches on the Anaerobic-Anoxic-Oxic (A2O) process have traditionally evaluated the removal of nutrients or organic compounds separately, while the limited information is available on how surfactants affect nutrient reduction in the A2O process. Therefore, the main objective of this study was to evaluate an A2O process for simultaneous removal of surfactants and nutrients ( N and P) from municipal wastewater, emphasis on the system efficiency and the limitation.

Materials and methods

Study location and sample collection

The studied municipal wastewater treatment plant (WWTP) is located in Yazd Province, Iran (32°24.24′N. 53°56.13′E). The plant employs a conventional treatment initiated by preliminary units (manual bar screen, coarse bar screen, fine bar screen, grit chamber) for the removal of large solids and inert materials. Subsequent flow equalization stabilizes hydraulic and organic loading, which is then directed to a biological nutrient removal sequence - anaerobic, anoxic, and oxic tanks—for the removal of carbon, nitrogen, and phosphorus. The process concludes with solid-liquid separation in a secondary clarifier tank and effluent disinfection via chlorination (Fig. 1).

Fig. 1.

Fig. 1

Open in a new tab

Schematic of the investigated A2O wastewater treatment plant (WWTP (SP, sampling points)).

Samples were collected monthly for one year, from September 23, 2023 to September 22, 2024. As shown in Fig. 1, the wastewater samples were taken from multiple points in the treatment process, including the plant influent, the influent and effluent of the anaerobic tank, and the anaerobic tank consists of two separate units: the first section is designed to receive and mix the returned activated sludge (RAS) and the second section acts as the biological phosphorus removal (BPR) zoned the effluents of anoxic, oxic, and secondary clarifier tanks.

Experimental procedures for wastewater quality measurement

One-liter polypropylene containers were used to collect grab samples. Then, the samples were immediately transferred to the laboratory to perform experiments. Figure 2 shows two examples of sample containers used in the present study.

Fig. 2.

Fig. 2

Open in a new tab

Sampling points for wastewater analysis.

The chemical oxygen demand (COD) digestion step was performed using a HACH digester (model DRB200, USA) at 150 °C for 2 h, with final readings recorded using a Lovibond photometer (model MD600, Italy). Biochemical oxygen demand (BOD), measurements were conducted using a Lovibond BOD meter (model BD600). Phosphorus was measured using the colorimetric method (ammonium molybdate reagent) and read with a spectrophotometer (Prove 300, Germany). Nitrate was also measured using the colorimetric method with 2-6-dimethylphenol reagent.

The linear alkylbenzene sulfonate (LAS) was measured using the methylene blue active substances (MBAS) method (Standard Method 5540-C). Initially, 100 mL of the sample was transferred into a separatory funnel, followed by the addition of 25 mL of methylene blue, 10 mL of chloroform, and 10 mL of isopropyl alcohol. The mixture was shaken for 30 s to allow the formation of two distinct phases: an organic phase (containing chloroform and dye) and an aqueous phase. The chloroform-containing solution, which appeared blue, was collected in a separate container. The separated aqueous phase was subsequently returned to the separatory funnel, and the extraction procedure was repeated twice, with 10 mL of chloroform added in each cycle. The collected chloroform solutions were combined in the separatory funnel, followed by the addition of 50 mL of washing solution, and the mixture was subsequently shaken vigorously for 30 s. The settled chloroform solution was then filtered, transferred into a 100 mL volumetric flask, and the volume adjusted to the mark. Finally, the sample absorbance was determined at a wavelength of 625 nm29.

This study was approved by the Ethics Committee of Shahid Sadoughi University of Medical Sciences (IR.SSU.SPH.REC.1403.140).

Data analysis

Statistical analysis was performed with a mixed linear model, and significant main effects were identified using Bonferroni-corrected pairwise comparisons, all conducted within the SPSS statistical software (version 27). A one-sample t-test was employed to compare the measured effluent concentrations against available standard values.

Results

Removal efficiency of organic parameters

The research initially characterized seasonal fluctuations in COD at throughout the treatment process. Sampling points included the raw wastewater influent, the anaerobic tanks influent and effluent streams, and the effluent discharges from the anoxic, oxic, and secondary clarifier unites. Figure 3 shows the COD fluctuations in sampling points.

Fig. 3.

Fig. 3

Open in a new tab

Seasonal variations of COD during the A2O process.

As shown in Fig. 3, the highest influent COD concentration was recorded in winter (578 ± 104.21 mg/L), while the lowest COD concentration was observed in summer (515.41 ± 52.26 mg/L). The chemical oxygen demand (COD) removal efficiency demonstrated seasonal variation, with values of 97.01% in autumn, 96.20% in winter, 96.05% in spring, and 96.07% in summer. The total COD removal efficiency was 96.35% in all of the study time. The results of statistical analysis showed that there was a significant difference in the COD removal between anaerobic - anoxic (P < 0.05) and anoxic - oxic (P < 0.05) tanks.

Figure 4 shows the BOD fluctuations in sampling points.

Fig. 4.

Fig. 4

Open in a new tab

Seasonal variations of BOD during the A2O process.

As shown in Fig. 4, the highest influent BOD concentration was in the autumn season (300.5 ± 57.24 mg/L) and the lowest BOD concentration was observed in summer (260.83 ± 22.61 mg/L). The biochemical oxygen demand (BOD) removal efficiency demonstrated seasonal variation, with values of 98.14% in autumn, 97.97% in winter, 97.4% in spring, and 97.18% in summer. The total BOD removal efficiency was 97.69% in all of the study period. There was a significant difference in the BOD removal efficiency between the anaerobic - anoxic (P < 0.05) and anoxic - oxic (P < 0.05) tanks.

Evaluation of nutrient removal efficiency

The performance of the different units in phosphorus removal is presented in Fig. 5.

Fig. 5.

Fig. 5

Open in a new tab

Seasonal variations of phosphorus during the A2O process.

As shown in Fig. 5, the highest influent phosphorus concentration was occurred in summer (6 ± 0.19 mg P/L), while the lowest was observed in winter (5.62 ± 0.42 mg P/L). The phosphorus removal efficiency demonstrated seasonal variation, with values of − 1.2% in autumn, 7.11% in winter, 5.29% in spring, and 12.33% in summer. The phosphorus removal efficiency was 23.49% in all of the study period. The significant difference observed between the anoxic - oxic tanks (P = 0.006).

Also, the performance of the different units in nitrate removal is presented in Fig. 6.

Fig. 6.

Fig. 6

Open in a new tab

Seasonal variations of nitrate during the A2O process.

As shown in Fig. 6, the highest nitrate concentration was occurred in summer (1.08 ± 0.13 mg NO₃–N /L as), while the lowest was observed in autumn (0.37 ± 0.07 mg NO₃–N /L). Seasonal analysis showed that the A2O process was accompanied by an increase in nitrate concentration in all seasons, indicating active nitrification in the system. Statistical results showed that there was a significant difference between anoxic and oxic tanks in nitrate removal (P < 0.001), indicating a relative reduction of nitrate in the anoxic tank.

Linear alkylbenzene sulfonate removal efficiency

A primary objective of the current study was to evaluate the efficacy of an A2O system for the concurrent removal of detergents and nutrients. Figure 7 shows the residual detergent (LAS) as the unit type in the A2O process.

Fig. 7.

Fig. 7

Open in a new tab

Seasonal variations of detergent (LAS) during the A2O process.

As shown, the highest detergent (LAS) concentration was observed in winter (5.53 ± 1 mg LAS/L), whereas the lowest occurred in autumn (2.41 ± 1.12 mg LAS/L). The efficacy of the A2O process in detergent removal was evaluated as seasonal variations. The detergent (LAS) removal efficiency demonstrated seasonal variation, with values of 92.11% in autumn, 95.84% in winter, 93.54% in spring, and 93.95% in summer. Statistical tests showed no significant difference in detergent removal among the tanks; in other words, the removal rate was the same in all tanks.

Table 2 compares the average parameters of the system final effluent with the standards published by the Iranian Environmental Protection Organization for wastewater discharge and reuse.

Table 2.

Comparison of final effluent quality with Iranian Standards30.

Parameters Values Unit Output standards
Discharge into
surface waters		 P value Discharge into
absorbent wells		 P value Irrigation P value
COD 20.12 ± 2.44 mg/L 60 < 0.001 60 < 0.001 200 < 0.001
BOD 6.64 ± 0.96 mg/L 30 < 0.001 30 < 0.001 100 < 0.001
Phosphorus 4.46 ± 0.18 mg-P/L 6 < 0.001 6 < 0.001 – –
Nitrate*

13.18 ± 3.7

(58.38 NO3)

 mg NO3-N/L 50 0.3 10 0.01 – –
Detergent 0.22 ± 0.05 mg-LAS/L 1.5 < 0.001 0.5 0.002 0.5 0.002
Open in a new tab

* The nitrate concentration in this study is reported as mg NO3- N/L, while the standard concentrations are expressed as mg NO₃/L).

The concentration of COD, BOD, phosphorus, and detergent (LAS) in the final effluent were significantly below the national standard limits. Nitrate was the only parameter that exceeded the permissible limit compared to the absorption well standard.

Discussion

Given the excessive use of detergents in domestic and industrial applications, and the presence of harmful nutrients in municipal wastewater (such as phosphorus and nitrogen), improving the efficiency of wastewater treatment plants is of special importance.

In the present work, one of the main parameters was COD. As data finding, there is a significant fluctuation in COD (from 465.5 to 700 mg/L), indicating qualitative variations in municipal wastewater (Fig. 3). These variations may be due to seasonal conditions, rainfall, or the influx of industrial wastewater into the municipal wastewater collection system. In contrast, the range of BOD fluctuations (232–366.25 mg/L) is considerably lower than that of COD (Fig. 4). This difference may be attributed to the occurrence of resistant and slowly biodegradable organic compounds that affect the COD concentration31. The preliminary treatment in the A2O process includes screening and grit removal units, which in this study were able to remove a relatively small portion of COD (6.1%) and BOD (8.68%). The purpose of designing these units is to remove heavy and inorganic particles; therefore, the removal observed in this study may be attributed to the elimination of organic matter adhered to suspended solids. According to reports by Wen et al., if the screening and grit removal units operate simultaneously and efficiently, COD removal will range between 5% and 15%, and BOD₅ removal between 3% and 10%. These units primarily reduce coarse and insoluble pollutants and have little effect on removing soluble pollutants32. The minimum and maximum removal efficiencies of COD and BOD in the anaerobic and oxic tanks were 6.11% and 85.78%, and 8.48% and 88.92%, respectively. As observed, the oxic tank exhibited the highest organic matter removal efficiency. These findings suggest that a combination of anaerobic, anoxic, and oxic processes along with secondary clarifier can effectively treat municipal wastewater with variable organic loads. The present study results showed that the difference between COD and BOD with standards was statistically significant. Rahimi et al. reported a COD removal efficiency of 94.5% while investigating the performance of the A2O system for industrial wastewater from dairy products33 that is in accordance with the present study.

The phosphorus concentration remained relatively constant in all units of the treatment plant (Fig. 5). Also, the phosphorus concentration in the influent wastewater was lower than the concentration observed in the influent to the anaerobic tank. Latter finding could be due to several factors. In the A2O process, waste activated sludge management is a critical factor governing the removal efficiency of nutrients, especially phosphorus. In the studied wastewater treatment plant, the excess sludge from the secondary clarifier unit is transferred to a sludge drying bed, due to anaerobic conditions and long retention time, phosphate accumulating bacteria (PAOs) release phosphate stored in their cells. As a result, the sludge drying bed effluent is enriched with soluble phosphate. This effluent is returned to the beginning of the anaerobic tank, thereby increasing the phosphorus load entering the system. These conditions have reduced the phosphorus removal efficiency in the early stages of the treatment and led to negative removal efficiency in this unit.

Phosphorus removal in the different treatment units, particularly in the anaerobic tank, has been limited due to unfavorable operating conditions. The limited phosphorus removal in the anaerobic tank may be related to insufficient nitrate removal. The returned sludge, which contains an amount of nitrate, is transferred to the anaerobic tank. In this condition, the tank will change from anaerobic to anoxic due to the oxygen present in the nitrate. The competition between phosphate accumulating bacteria (PAOs) and denitrifying bacteria favors the latter, reducing phosphorus removal efficiency3,34. Phosphorus removal efficiency varied by season, with the highest efficiency observed in the oxic tank during summer (Fig. 5). This finding may be related from increased enzymatic activity of phosphate-accumulating organisms (PAOs). These bacteria perform optimally at temperatures between 20 and 30 °C, while higher temperatures cause their gradual decline in the system35.

Also, the characteristics of the influent wastewater to the system can be influential. Lu and Shim, investigated the effect of chloride on the A2O process performance. They found that increasing the chloride ion concentration from 150 to 500 mg/L led to increase the COD (15 to 25 mg/L) and ammonia (0.05 to 0.08 mg/L) levels in the effluent, while total phosphorus and total nitrogen concentrations remained constant. However, when the chloride concentration exceeded 1000 mg/L, both total phosphorus (0.8 to 0.9 mg/L) and total nitrogen (2.4 to 2.5 mg/L) in the effluent increased as well36. Xiao et al. achieved a phosphorus removal efficiency of 97% (from 8.7 to 0.32 mg P/L) in municipal wastewater treatment using the A2O process37. Rong et al., in a similar study, reported a removal efficiency of 93.6%38. These reported phosphorus removal efficiencies are considerably higher than those obtained in the present study. Based on the studies conducted, it can be concluded that the A2O process has the potential to achieve high phosphorus removal efficiency. Therefore, it is necessary to optimize the operating conditions in the system under study. The mean phosphorus concentration in the effluent differed significantly from the standard limit.

The results showed that the nitrate concentration was very low (0.6 ± 0.32 mg NO₃–N /L) in the wastewater influent, which is normal (Fig. 6). In raw wastewater, nitrogen is not present as nitrate or nitrite. Nitrogen occurs in both organic and ammonia forms. In the influent to the anaerobic tank, a slight increase in nitrate (1.34 ± 0.72 mg NO₃-N /L) was observed, likely due to partial oxidation of ammonia nitrogen and the presence of nitrate in the sludge drying bed effluent. In the anaerobic tank, the nitrate concentration (4.54 ± 2.65 mg NO3-N /L) increased, which could be due to limited nitrogen oxidation under low oxygen conditions (during this study, the average dissolved oxygen concentration in the anaerobic tank was 0.23 mg/L). In the anoxic tank, the nitrate concentration decreases to 2.45 ± 2.03 mg NO₃–N /L, indicating a relatively good performance of the denitrification process (dissolved oxygen concentration in the anoxic tank was less than 0.5 mg/L). In the oxic tank, with the presence of oxygen and the activity of nitrification bacteria, ammonia nitrogen is oxidized and converted into nitrite, followed by nitrate production. Nitrate concentrations increased significantly, which may be attributed to the average dissolved oxygen concentration of 4.8 mg/L maintained in the oxic tank during the study period. Increasing dissolved oxygen will increase the activity of nitrification bacteria and subsequently lead to increase nitrate production in the oxic tank. As a result, the nitrate load in the anoxic tank will be high. Tian et al. achieved a removal efficiency of 27% for nitrate in municipal wastewater treatment with a full-scale A2O process39. Nitrate concentration in the clarifier effluent has been associated with an increase, which could be due to the release of some nitrate stored in the cells or the conversion of residual ammonia to nitrate by Nitrobacter and Nitrosomonas bacteria active in the clarifier. Statistical analysis showed that the difference in nitrate concentration was not significant compared to the standard for discharge to surface water, but no significant difference was observed compared to the standard for discharge to absorption wells and agricultural uses.

According to the detergent finding, the detergent concentration decreased significantly in the effluent of WWTP, indicating the effective removal of detergent by the A2O process. The highest detergent removal efficiency was related to the winter season (95.84%), and the lowest efficiency was related to the autumn season (92.11%). The A2O process has been very suitable and efficient for detergent removal. The results showed that the difference between the detergent (LAS) and the standards was statistically significant. In this study, the lowest detergent removal efficiency during the year occurred in autumn. Additionally, the highest organic load input was recorded in autumn. This correlation may suggest a detrimental impact of input organic load on the rate of surfactant removal.

In this study, the lowest detergent (LAS) removal efficiency throughout the year occurred in autumn. Additionally, the highest organic load input was recorded in autumn. This correlation could suggest a detrimental impact of input organic load on the rate of detergent removal. Seyed Salehi et al. evaluated the performance of a Moving Bed Sequential Continuous Flow Reactor (MSCR) for removing Linear Alkylbenzene Sulfonate (LAS) from municipal wastewater at a full scale. The maximum removal efficiency achieved at a hydraulic retention time of 2.5 h, with an input organic load of 300 mg/L and an input LAS of 15 mg/L, was 96.4%. At the same hydraulic retention time, with input organic loads of 500 mg/L and input LAS concentrations of 15 mg/L and 25 mg/L, the maximum removal efficiencies were 88.7% and 92.2%, respectively. The study revealed that the LAS removal efficiency decreased as the input organic load increased40. In another study, Nie et al. reported that the existence of LAS at a concentration of 5 mg/L in an anaerobic membrane bioreactor process led to reduced COD removal (95.2%) and decreased biogas production compared to the control samples (96.8%)41.

Regarding the removal mechanism of linear alkylbenzene sulfonate (LAS) in the A2O system, both adsorption on activated sludge and microbial biodegradation may occur. However, biodegradation is the dominant elimination pathway. LAS are an anionic surfactant that is rapidly biodegradable, especially under aerobic conditions, where heterotrophic microorganisms play a major role in its degradation. Kaida et al. reported that among the removal mechanisms, only about 3–15% of LAS is temporarily removed through adsorption onto activated sludge, while 93–97% is removed through biological degradation, mainly under aerobic conditions. Since LAS does not degrade well under anaerobic or anoxic conditions but degrades rapidly in the aerobic zone, its accumulation in sludge is generally limited42.

In this study, although detergents were effectively removed, phosphorus removal remained unsatisfactory. This occurrence can be attributed to one of the following factors: First, detergents may harm the microorganisms present in biological treatment systems, especially in the anaerobic and oxic units. Detergents have different antibacterial properties due to their characteristics and can limit the metabolic activities and growth rate of aquatic bacteria. Detergents inhibit cellular metabolism by destroying proteins in the cytoplasmic membrane of the bacterial cell. When the detergent concentration surpasses the critical limit, the bacterial cell membrane can disintegrate entirely. The presence of detergents may lead to poisoning of microorganisms and reduce their ability to remove phosphorus effectively43. In general, ionic detergents are more toxic than nonionic detergents. Mitru et al. reported that high concentrations of anionic detergent (> 25 mg/L) in wastewater treatment plants can impair performance and efficiency due to foam formation and biological toxicity. Detergent concentrations greater than or equal to 100 mg/L in activated sludge systems also inhibit the sludge degradation capacity17. Secondly, detergents contain very low percentages of phosphates or are phosphate-free in many wastewater systems in all of the worlds. Over time, compounds such as zeolite, poly-carboxylic acids, and sodium carbonate have replaced sodium tripolyphosphate (Na₅P₃O₁₀) compounds44. As a result, properly removing detergent in the wastewater treatment plant will not necessarily lead to the effective removal of phosphorus. Bering et al. reported efficiencies of 98.8% for anionic surfactants and 99.4% for nonionic surfactants in treating industrial laundry wastewater using the MBR process43. In another study, Kruszelnicka et al. achieved a 98% removal efficiency of anionic surfactants using the wetland process, and an 88% efficiency using the SBR process in domestic wastewater treatment45.

Gomes de Quevedo et al. reported that detergent products available on the market have low phosphorus concentrations, with a maximum of 0.01% by weight. Therefore, the contribution of detergents to the total phosphorus content in the wastewater of the studied treatment plants is negligible, with an estimated value of approximately 0.0014 g/person/day. This amount is equivalent to about 0.17% of the total phosphorus content in the wastewater46. Chen et al. reported that total detergent consumption tripled from 2000 to 2018. However, the phosphorus input to surface waters from detergents decreased by 35% during the same period47. The findings show that the average concentration of total phosphorus in wastewater streams has decreased in recent years and is now in the range of 3.5–6.7 mg/L. Also, the fraction of organic phosphorus accounts for an average of about 64% of total phosphorus, which is higher than the fraction of inorganic phosphorus. This issue indicates a change in the phosphorus composition of wastewater compared to the past, which is probably caused by the decrease in the amount of phosphate in the formulation of detergents48.

Conclusion

The primary objective of this study was to evaluate the co-removal efficiencies of detergents and nutrients in a wastewater treatment system. The results demonstrate that under optimal conditions, the Anaerobic-Anoxic-Oxic (A2O) process is an effective and sustainable method for the simultaneous removal of detergents, organic matter, and nutrients from industrial and municipal wastewater. The process achieved high removal efficiencies for COD and BOD, indicating excellent microbial activity in the oxic tank and an effective integration of the anaerobic, anoxic, and oxic stages.

However, phosphorus removal efficiency was suboptimal. This limitation could be attributed to three primary factors includes: the recycle of phosphorus from the sludge drying bed to the anaerobic tank, the presence of nitrate in the return sludge stream, which can impair biological phosphorus release, and also the potential inhibitory effects of detergents on the polyphosphate-accumulating organisms (PAOs) responsible for enhanced biological phosphorus removal. Reducing aeration time as a phosphorus control strategy decreases nitrate production and lowers nitrate concentration in the returned sludge, preventing excess nitrate from entering the anaerobic tank.

Acknowledgements

The authors thank Shahid Sadoughi University of Medical Sciences.

Author contributions

Fahimeh Teimouri and Tahereh Zarei Mahmoudabadi designed the study. Sampling and experiments were performed by Tahereh Zarei Mahmoudabadi and Amir Hossein Bagheri. Data analysis was conducted by Sara Jambarsang, and data interpretation was performed by Fahimeh Teimouri. The first draft of the manuscript was written by Tahereh Zarei Mahmoudabadi, and the finalization of the manuscript was carried out by Fahimeh Teimouri. Supervision was the responsibility of Fahimeh Teimouri. These are the authors’ roles.

Funding

No funding was received for this manuscript.

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  • 1.Brownlie, W. J. et al. Phosphorus price spikes: A wake-up call for phosphorus resilience. Front. Sustain. Food Syst.10.3389/fsufs.2023.1088776 (2023). [Google Scholar]
  • 2.Gao, L. et al. Simultaneous nitrate and dissolved organic matter removal from wastewater treatment plant effluent in a solid-phase denitrification biofilm reactor. Bioresour. Technol.314, 123714 (2020). [DOI] [PubMed] [Google Scholar]
  • 3.Velusamy, K. et al. Advanced techniques to remove phosphates and nitrates from waters: A review. Environ. Chem. Lett.19(4), 3165–3180 (2021). [Google Scholar]
  • 4.Sevda, S., Sreekishnan, T. R., Pous, N., Puig, S. & Pant, D. Bioelectroremediation of perchlorate and nitrate contaminated water: A review. Bioresour. Technol.255, 331–339 (2018). [DOI] [PubMed] [Google Scholar]
  • 5.Ward, M. H. et al. Drinking water nitrate and human health: An updated review. Int. J. Environ. Res. Public Health15(7), 155 (2018). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Lin, L. et al. Nitrate contamination in drinking water and adverse reproductive and birth outcomes: A systematic review and meta-analysis. Sci. Rep.13(1), 563 (2023). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Shaban, J., Al-Najar, H., Kocadal, K., Almghari, K. & Saygi, S. The effect of nitrate-contaminated drinking water and vegetables on the prevalence of acquired methemoglobinemia in Beit Lahia City in Palestine. Water15(11), 1989 (2023). [Google Scholar]
  • 8.Wu, B., Wan, J., Zhang, Y., Pan, B. & Lo, I. M. Selective phosphate removal from water and wastewater using sorption: Process fundamentals and removal mechanisms. Environ. Sci. Technol.54(1), 50–66 (2019). [DOI] [PubMed] [Google Scholar]
  • 9.Usman, M. O., Aturagaba, G., Ntale, M. & Nyakairu, G. W. A review of adsorption techniques for removal of phosphates from wastewater. Water Sci. Technol.86(12), 3113–3132 (2022). [DOI] [PubMed] [Google Scholar]
  • 10.Venkiteshwaran, K., McNamara, P. J. & Mayer, B. K. Meta-analysis of non-reactive phosphorus in water, wastewater, and sludge, and strategies to convert it for enhanced phosphorus removal and recovery. Sci. Total Environ.644, 661–674 (2018). [DOI] [PubMed] [Google Scholar]
  • 11.Erem, S. & Razzaque, M. S. Dietary phosphate toxicity: An emerging global health concern. Histochem. Cell Biol.150(6), 711–719 (2018). [DOI] [PubMed] [Google Scholar]
  • 12.Razavi, T., Fadaei, A., Sadeghi, M. & Sedehi, M. Evaluation of the photosonolysis process efficacy for the removal of anionic surfactant linear alkyl benzene sulfonate from aqueous solutions. J. Ecol. Eng.21(6), 1–7 (2020). [Google Scholar]
  • 13.Mohammadi, V., Tabatabaee, M., Fadaei, A., Mirhoseini, S.A.: Comparing activated carbon and magnetic activated carbon in removal of linear alkylbenzene sulfonate from aqueous solution by heterogeneous catalytic ozonation process. Iran. J. Catal.10(3), (2020).
  • 14.Villabona-Ortíz, A., Tejada-Tovar, C., De-La-Rosa-Jiménez., L. Evaluation of parameters in the removal of linear alkylbenzene sulfonate anionic surfactant using electrocoagulation. Rev. Fac. Ing.30(55), (2021).
  • 15.Mondal, B., Adak, A. & Datta, P. Integrated UV–H2O2 and biological treatment processes for the removal of cationic surfactant. J. Environ. Eng. Sci.16(2), 85–93 (2021). [Google Scholar]
  • 16.Tu, R. et al. Treatment of wastewater containing linear alkylbenzene sulfonate by bacterial-microalgal biological turntable. Korean J. Chem. Eng.37(5), 827–834 (2020). [Google Scholar]
  • 17.Mitru, D. et al. Removal and effects of surfactants in activated sludge system. Rev. Chim.71(1), 100–106 (2020). [Google Scholar]
  • 18.Mojab, S., Moussavi, S. G. & Shekoohiyan, S. The investigation of biological degradation of sodium dodecyl sulfate and its associated COD in wastewater using anoxic baffled reactor. Iran. J. Health Environ.18(1), 1–24 (2025). [Google Scholar]
  • 19.Mondal, B., Adak, A. & Datta, P. Degradation of anionic surfactant in municipal wastewater by UV-H2O2: Process optimization using response surface methodology. J. Photochem. Photobiol. A. Chem.375, 237–243 (2019). [Google Scholar]
  • 20.Klimonda, A. & Kowalska, I. From environmental threat to control: A review of technologies for removal of quaternary ammonium compounds from wastewater. Membranes16(1), 1 (2026). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Wang, X. et al. Environmental concentrations of surfactants as a trigger for climax of horizonal gene transfer of antibiotic resistance. Heliyon9(6), e17034 (2023). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Kulkarni, D., Jaspal, D. & Itankar, N. Surfactant removal from wastewater: A comparative review on adsorption versus other techniques. Environ. Technol. Rev.14(1), 713–742 (2025). [Google Scholar]
  • 23.Tang, G. et al. Variation of effluent organic matter (EfOM) during anaerobic/anoxic/oxic (A2O) wastewater treatment processes. Water Res.178, 115830 (2020). [DOI] [PubMed] [Google Scholar]
  • 24.Han, Y., Yang, K., Yang, T., Zhang, M. & Li, L. Bioaerosols emission and exposure risk of a wastewater treatment plant with A2O treatment process. Ecotoxicol. Environ. Saf.169, 161–168 (2019). [DOI] [PubMed] [Google Scholar]
  • 25.Rabani Mehr, A., Nikkhah, H., Farhadian, M. Techno-economic analysis of a municipal wastewater treatment plant: A comparison among SBR, MLE & A2O Processes. Sci. Iran. (2023).
  • 26.Liang, J., Yuan, Y., Zhang, Z., You, S. & Yuan, Y. Modeling a three-stage biological trickling filter based on the A2O process for sewage treatment. Water13(9), 1152 (2020). [Google Scholar]
  • 27.Villanueva, J. A. L., Tulipan, J. U., Detras, M. C. M. & Ventura, J.-R.S. Effect of C/N ratio on COD and nitrite removal in plastic recycling wastewater treatment using the anaerobic-anoxic-oxic (A2O) system. J. Indian Chem. Soc.102(12), 102270 (2025). [Google Scholar]
  • 28.Zhou, Y., Zhu, J., Huang, X., Gao, J. & Zhuang, G. Performance and microbial characteristics of anaerobic/anoxic/aerobic (A2O)-biofilm system for electroplating wastewater treatment: A pilot scale study. Desalin. Water Treat.313, 74–81 (2023). [Google Scholar]
  • 29.Lipps, W.C., Braun-Howland, E.B., Baxter, T.E. Standard methods for the examination of water and wastewater. 24th edn. 1536 (American Public Health Association, American Water Works Association, Washington DC, 2023).
  • 30.Department of Environment (DOE). Iranian environmental quality standards for effluent discharge (DOE, 2012). [Google Scholar]
  • 31.Tabraiz, S. et al. Effect of effluent and sludge recirculation ratios on integrated fixed films A2O system nutrients removal efficiency treating sewage. Desal. Water Treat.114, 120–127 (2018). [Google Scholar]
  • 32.Wen, H. et al. Investigation of organic content and microbial characteristics distribution at the pretreatment units for wastewater treatment plant sustainable upgrading. Desal. Water Treat.320, 100660 (2024). [Google Scholar]
  • 33.Rahimi, Z., Zinatizadeh, A. A., Zinadini, S., van Loosdrecht, M. & Batstone, D. J. Concurrent removal of carbon and nutrients in a one-stage dual internal circulation airlift A2O bioreactor from milk processing industrial wastewater: Process optimization, sludge characteristics and operating cost evaluation. Chemosphere355, 141804 (2024). [DOI] [PubMed] [Google Scholar]
  • 34.Zhang, W., Huang, F. & Hu, W. Performance and mechanism of synchronous nitrate and phosphorus removal in constructed pyrite-based mixotrophic denitrification system from secondary effluent. Environ. Sci. Pollut. Res.27(29), 36816–36825 (2020). [DOI] [PubMed] [Google Scholar]
  • 35.Izadi, P., Izadi, P. & Eldyasti, A. Design, operation and technology configurations for enhanced biological phosphorus removal (EBPR) process: A review. Rev. Environ. Sci. Biotechnol.19(3), 561–593 (2020). [Google Scholar]
  • 36.Lu, Q. & Shim, H. Comparison of chloride effect between A2O and SBR processes treating domestic wastewater. Desal. Water Treat.53(9), 2549–2554 (2015). [Google Scholar]
  • 37.Xiao, K. et al. Nitrogen and phosphorus removal using fluidized-carriers in a full-scale A2O biofilm system. Biochem. Eng. J.115, 47–55 (2016). [Google Scholar]
  • 38.Rong, Y. et al. Advanced nutrient removal in a continuous A2O process based on partial nitrification-anammox and denitrifying phosphorus removal. J. Water Process. Eng.36, 101245 (2023). [Google Scholar]
  • 39.Tian, M. et al. The first metagenome of activated sludge from full-scale anaerobic/anoxic/oxic (A2O) nitrogen and phosphorus removal reactor using Illumina sequencing. J. Environ. Sci.35, 181–190 (2015). [DOI] [PubMed] [Google Scholar]
  • 40.Seyedsalehi, M. et al. Performance evaluation of several sequencing batch biofilm reactors with movable bed in treatment of linear alkyl benzene sulfonate in urban wastewater. Int. J. Environ. Sci. Technol.16(11), 6763–6772 (2019). [Google Scholar]
  • 41.Nie, Y. et al. Effect of anionic surfactant inhibition on sewage treatment by a submerged anaerobic membrane bioreactor: Efficiency, sludge activity and methane recovery. Chem. Eng. J.315, 83–91 (2017). [Google Scholar]
  • 42.Kaida, H., Syed, M. A., Shukor, M. Y. & Othman, A. R. Biodegradation of linear alkylbenzene sulfonates (LAS): A mini review. Bioremed. Sci. Technol. Res.9(1), 1–6 (2021). [Google Scholar]
  • 43.Bering, S. et al. Removal of organic pollutants and surfactants from laundry wastewater in membrane bioreactor (MBR). Desalin. Water Treat.134, 281–288 (2018). [Google Scholar]
  • 44.Quevedo, C. M. G., Piveli, R. P. & Paganini, W. D. S. Influence of the detergent formulation on the concentration of phosphorus in the sewage inflows to the WWTPs: The Brazilian experience. Environ. Technol.39(10), 1260–1270 (2018). [DOI] [PubMed] [Google Scholar]
  • 45.Kruszelnicka, I., Ginter-Kramarczyk, D., Wyrwas, B. & Idkowiak, J. Evaluation of surfactant removal efficiency in selected domestic wastewater treatment plants in Poland. J. Environ. Health Sci. Eng.17(2), 1257–1264 (2019). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 46.Gomes de Quevedo, C. M. & da Silva Paganini, W. Detergents as a source of phosphorus in sewage: The current situation in Brazil. Water Air Soil Pollut.227(1), 14 (2015). [Google Scholar]
  • 47.Chen, X. et al. Mitigating phosphorus pollution from detergents in the surface waters of China. Sci. Total Environ.804, 150125 (2022). [DOI] [PubMed] [Google Scholar]
  • 48.Bischak, E., Ghezzehei, T. A. & Ryals, R. Nitrogen and phosphorus mineralization dynamics in human excreta-derived fertilizers. Front. Agron.10.3389/fagro.2024.1425461 (2024). [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Articles from Scientific Reports are provided here courtesy of Nature Publishing Group

RESOURCES