Wastewater treatment is one of the most important factors of controlling pollution in the environment (EPA).  Most treatment plants are built to purify water to be discharged back into streams and other bodies of water (Environmental Protection Agency, 1998).  Population and industrial growth have increased the demand on our natural resources (USEPA, 2004). Progress in reducing pollution has barely kept ahead of population growth, changes in industrial processes, technological developments, changes in land use, business innovations, and many other factors (USEPA, 2004). Increases in both the quantity and variety of goods produced can greatly alter the amount and complexity of industrial wastes and challenge traditional treatment technology (Environmental Protection Agency, 1998). The application of commercial fertilizers and pesticides, combined with sediment from growing development activities, continues to be a source of significant pollution as runoff washes off the land (USEPA, 2004).

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In recent decades, one of the main issues in aquatic environments is the emerging problem of micropollutants such as pharmaceuticals, personal care products, hormones, detergents, and disinfectants due to their potential risk on the ecosystem (Xu et al., 2016). Thousands of tons of pharmaceuticals are used annually in human and veterinary medicine to prevent illness, as well as growth promoters in livestock and fish, and in agriculture (Xu et al., 2016). After being administrated, pharmaceuticals could be excreted in unchanged forms or metabolites in urine and faeces, along with direct disposal from manufactures as sources into the environment (Xu et al., 2016).

Current wastewater treatment processes have three stages: physical (primary treatment), biological (secondary treatment), and a chemical (coagulation sedimentation).  The primary treatment is responsible for removing the solids from wastewater by passing through screens and using gravity to settle the solids out of the water.   The secondary treatment addition of oxygen to wastewater, masses of microorganisms grew and rapidly metabolized organic pollutants.  The coagulation sedimentation uses chemicals such as alum, lime or iron salts which are added to wastewater to cause certain pollutants, such as phosphorus, to bunch together into large, heavier masses which can be removed faster through physical processes.  It does this by alum reacting with phosphoric acid and producing aluminum phosphate which is a solid (Zhao et al., 2015). 

Carbon, nitrogen, and phosphorus are essential to living organisms and are the main nutrients present in natural water (USEPA, 2004). Large amounts of these nutrients are also present in sewage, some industrial wastes, and drainage from fertilized land (USEPA, 2004).

Current secondary biological treatment processes do not remove the phosphorus and nitrogen to any substantial extent, and they may actually convert the organic forms of these substances into mineral form, making them more usable by plant life (USEPA, 2004). The release of large amounts of nutrients, primarily phosphorus but occasionally nitrogen, causes nutrient enrichment which results in excessive growth of algae, which can lead to eutrophication (Ward et al, 2018).  Uncontrolled algae growth blocks out sunlight and chokes aquatic plants and animals by depleting dissolved oxygen in the water at night (USEPA, 2004).

By providing additional biological treatment beyond the secondary stage, nitrifying bacteria present in wastewater treatment can biologically convert ammonia to the non-toxic nitrate through a process known as nitrification (USEPA, 2004). The nitrification process is normally sufficient to remove the toxicity associated with ammonia in the effluent (USEPA, 2004). Since nitrate is also a nutrient, excess amounts can contribute to the uncontrolled growth of algae. However, excessive concentrations of nitrate in drinking water can also be hazardous to health, especially for infants and pregnant women. The maximum contaminant level (MCL) for nitrate in public drinking water supplies in the United States is 10 mg/L (Ward et al., 2018).  The MCL was set to protect against infant methemoglobinemia (MetHb), which results in blue baby syndrome.  Through endogenous nitrosation, nitrate is a starting compound in the formation of N-nitroso compounds (NOC), and most NOC’s are carcinogens and teratogens (Bhatnagar & Sillanpää, 2011). Therefore, exposure to NOC’s formed after ingestion of nitrate from drinking water and dietary sources may result in cancer, birth defects, or other adverse health effects (Bhatnagar & Sillanpää, 2011).

Wastewater treatment plants (WWTP) are one of the main ways that pharmaceuticals enter into the environment.  This is because there is incomplete removal for most compounds because WWTPs were not designed to remove pharmaceuticals (Quintana et al., 2016).  Although the direct and indirect effects of pharmaceuticals in aquatic systems are not yet well known, studies done in labs exemplify the potential adverse effects (Dawas-Massalha et al., 2014). These effects include how the spread of antibiotics can accelerate the evolutionary resistance in naturally occurring bacteria; and some drugs have the potential to accumulate in soils and can lead to being adsorbed into crops during irrigation with recycled water (Dawas-Massalha et al., 2014). 

Xu et al. (2016) showed that wastewater treatment processes with significant nitrification, tend to have higher removal rates of pharmaceuticals than those without nitrification, especially for 17α-ethinylestradiol (EE2).  It was also demonstrated that the effluent concentration and removal of ammonium showed significant correlation with overall target pharmaceutical removal (Xu et al., 2016).  The nitrification process was reported to enhance pharmaceutical removal through cometabolic biodegradation by the ammonia oxidizing bacteria (AOB) (Xu et al., 2016).  The AOB’s non-specific enzyme ammonia monooxygenase (AMO), is able to alter aliphatic and aromatic compounds, which can to help remove pharmaceuticals by AOB (Xu et al., 2016).  This is because during the nitrification process, AOB converts NH4 + to NO2−, which degrades a broad range of aromatic compounds through AMO, such as hydrocarbons, phenol, and aromatic compounds (Xu et al., 2016).

Pharmaceuticals could be biodegraded by microorganisms in the presence of a growth substrate, such as easily degradable compounds or nutrients (Quintana et al., 2005). Bezafibrate, naproxen, and ibuprofen showed different degrees of conversion and mineralization in the presence of an external carbon source, while they were not degraded metabolically (Quintana et al., 2005). The biodegradation of pharmaceuticals by AOB were shown to follow the cometabolic pathway, with ammonia being used as the primary substrate and energy source for microbial growth and enzyme introduction (Xu et al.,2016).

The active site of AMO contains metal ions such as copper ions (Xu et al.,2016). Oxygen reacts to convert Cu+-Cu+ into Cu2+-Cu2+ under aerobic conditions while the oxygen remains bound as an electrophilic radical (Xu et al.,2016). The oxygenated form of AMO will react with EE2 to produce Cu2+-Cu2+ and the biotransformation products (Xu et al.,2016). However, this model was proposed based on monooxygenase; the dioxygenase may also need to be considered in the pharmaceuticals biodegradation mechanism because this enzyme is used in the cometabolic or metabolic biodegradation, especially in mixed cultures consisting of different bacterial species (Xu et al.,2016).

AOB is able to convert pharmaceuticals under ammonium starvation conditions (Xu et al., 2016). The high concentration of ammonium inhibited the cometabolic biotransformation until the ammonia was depleted, and then the catabolic repression mechanism for cometabolism was activated (Dawas-Massalha et al., 2014). Therefore, in order to obtain maximum cometabolic degradation rate of pharmaceuticals, the concentration ratio between pharmaceuticals and ammonia should be maintained within a certain range ((Dawas-Massalha et al., 2014). Hydroxylation was another biodegradation pathway of AOB for pharmaceuticals including ibuprofen, trimethoprim, atenolol, and bezafibrate (Quintana et al., 2005). The enzyme AMO may be able to hydroxylate the pharmaceuticals, similar to how it does with ammonia in AOB (Quintana et al., 2005).

The biodegradation pathways and products for several pharmaceuticals are influenced by their structures (Xu et al., 2016). The incomplete biotransformation of pharmaceuticals in wastewater treatment processes could result in more toxic products being released into the water bodies (Dawas-Massalha et al., 2014).  The cometabolism by AOB could induce different biodegradation pathways of pharmaceuticals (Xu et al., 2016).  An example of this is that iopromide could be transformed into different products depending on if nitrification is involved (Xu et al., 2016).

In conclusion, the ammonia oxidizing bacteria (AOB) could significantly improve pharmaceutical removal due to the non-specific substrate range of the enzyme ammonia monooxygenase in AOB (Xu et al., 2016).  The pharmaceutical degradation is correlated to the nitrification rate in wastewater treatment.  The cometabolic biodegradation by AOB has been shown to have greater pharmaceutical removal than metabolic biodegradation (Xu et al., 2016).  The cometabolic biodegradation by AOB could involve reactions including oxidation and hydroxylation during nitrification, this could result in the formation of different products compared to the metabolic biodegradation (Xu et al., 2016). Further research is required to gain a better understanding of pharmaceuticals biotransformation by AOB. Ecotoxicological testing is still needed to be done to determine what a sufficient pharmaceutical concentration threshold; this is because there have been studies where even a 90% removal rate could be a cause for concern, which could induce adverse ecotoxicological effects on aquatic organisms (Dawas-Massalha et al., 2014).

References

Bhatnagar, A., & Sillanpää, M. (2011). A review of emerging adsorbents for nitrate removal from water. Chemical Engineering Journal, 168(2), 493-504.

Dawas-Massalha, A., Gur-Reznik, S., Lerman, S., Sabbah, I., & Dosoretz, C. G. (2014). Co-metabolic oxidation of pharmaceutical compounds by a nitrifying bacterial enrichment. Bioresource technology, 167, 336-342.

Environmental Protection Agency. (1998). How Wastewater Treatment Works The Basics. Retrieved from https://www3.epa.gov/npdes/pubs/bastre.pdf

Quintana, J. B., Weiss, S., & Reemtsma, T. (2005). Pathways and metabolites of microbial degradation of selected acidic pharmaceutical and their occurrence in municipal wastewater treated by a membrane bioreactor. Water research, 39(12), 2654-2664.

USEPA. (2004). Primer for Municipal Wastewater Treatment Systems.

Ward, M., Jones, R., Brender, J., de Kok, T., Weyer, P., Nolan, B., … & van Breda, S. (2018). Drinking Water Nitrate and Human Health: An Updated Review. International journal of environmental research and public health, 15(7), 1557.

Xu, Y., Yuan, Z., & Ni, B. J. (2016). Biotransformation of pharmaceuticals by ammonia oxidizing bacteria in wastewater treatment processes. Science of The Total Environment, 566, 796-805.

Zhao, Xiaohong, Zhao, Yaqian, Wang, Wenke, Yang, Yongzhe, Babatunde, Akintunde, Hu, Yuansheng, & Kumar, Lordwin. (2015). Key issues to consider when using alum sludge as substrate in constructed wetland. Water Science and Technology : A Journal of the International Association on Water Pollution Research, 71(12), 1775-82