The presence of anthropogenic contaminants, known as emerging pollutants (EPs), poses a major challenge for wastewater treatment. In line with the European Union’s vision for water recycling, efforts have focused on developing more sustainable advanced treatment technologies. To achieve this, circular economy principles can be applied by repurposing waste from other sectors. Consequently, drinking water treatment residuals (DWTR), by-products of drinking water treatment plants (DWTP), have emerged as promising, low-cost adsorbents for the removal of EPs in wastewater treatment. This study assessed the adsorption, in aqueous solution, of a mixture of three steroid hormones: estrone (E1), 17β-estradiol (E2), and 17α-ethinylestradiol (EE2), while also providing proof-of-concept insights using real wastewater. The adsorption process was evaluated by analysing the underlying mechanisms involved, with a focus on dosage, kinetics, and isotherm adsorption studies. The results showed that DWTR reached a maximum adsorption capacity of approximately 4 mg EPs/g DWTR within 24 h, suggesting potential compatibility with WWTP hydraulic retention times. In real wastewater (0.5 µg/L of E1, E2, and EE2), removal efficiency exceeded 50

Sample pretreatment is one of the most important steps in guaranteeing the success of a chromatographic analysis. The selected methodology must ensure simultaneously that a sample is “clean” enough for analysis and that the target analytes are not removed in the process. This can be especially difficult when working with complex matrices such as natural waters and wastewater. For pharmaceutical active compounds (PhACs) analysis by solid-phase extraction (SPE) followed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS), and due to the high level of organic matter in wastewater, the water samples are filtered consecutively through three filters, a paper filter, a glass microfiber filter of 1 µm, and a Nylon filter of 0.45 µm. This filtration allows the sample’s passage through the SPE cartridge to be faster, and there is no cartridge clogging, allowing for greater efficiency in the adsorption process. The big question is whether the PhACs are eliminated during filtration, since they may be adsorbed to organic matter. This work aimed to determine if the best approach for quantifying PhACs in wastewater and surface waters would be to filter them prior or to perform SPE directly. Both approaches analyzed a total of 26 PhACs. Turbidity (TUR) and permanganate index (PI) were determined, and their values were high for samples with a high organic matter content. A statistical analysis was performed to determine the best approach to treat these water samples and whether any correlation existed between PhAC concentrations, PI, and TUR. The PhAC quantification shows a positive correlation with TUR and a negative correlation with PI for most of the target PhACs. However, there are not significantly different results for filtered and not-filtered wastewater samples.

A unique cytochrome P450 (CYP) oxidoreductase (CPR) sustains activities of human microsomal CYPs. Its function requires toggling between a closed conformation enabling electron transfers from NADPH to FAD and then FMN cofactors and open conformations forming complexes and transferring electrons to CYPs. We previously demonstrated that distinct features of the hinge region linking the FAD and FMN domain (FD) modulate conformer poses and their interactions with CYPs. Specific FD residues contribute in a CYP isoform-dependent manner to the recognition and electron transfer mechanisms that are additionally modulated by the structure of CYP-bound substrate. To obtain insights into the underlying mechanisms, we analyzed how hinge region and FD mutations influence CYP1A2-mediated caffeine metabolism. Activities, metabolite profiles, regiospecificity and coupling efficiencies were evaluated in regard to the structural features and molecular dynamics of complexes bearing alternate substrate poses at the CYP active site. Studies reveal that FD variants not only modulate CYP activities but surprisingly the regiospecificity of reactions. Computational approaches evidenced that the considered mutations are generally in close contact with residues at the FD–CYP interface, exhibiting induced fits during complexation and modified dynamics depending on caffeine presence and orientation. It was concluded that dynamic coupling between FD mutations, the complex interface and CYP active site exist consistently with the observed regiospecific alterations.

The control of pharmaceutical compounds (PhCs) in urban wastewater treatment plants (WWTPs) is an issue of health and environmental relevance, expected to become mandatory in the European Union and beyond, but studies at full scale are still scarce. A long-term (2.5-year) full-scale study addressing practical strategies for controlling PhCs was conducted in two activated sludge WWTPs with nitrification (BEI with A2O process; FNW with oxidation ditch). The results showed similar removal patterns in both plants - some of the most abundant influent PhCs were highly removed, others presented intermediate and variable removals, and some were recalcitrant. These behaviours validated a 4-class (A-D) PhC framework (based on biodegradation (kbio) and solid-water distribution (Kd)) for interpreting and predicting PhC removal, particularly sensitive to kbio, with a turning point at 1 L/(gSS.d). A statistical analysis conducted to unveil the key operation variables showed T254 is a good indicator of PhC removal and confirmed that favouring conditions for nitrifiers to grow is an operational strategy to improve PhC control. In BEI, F/M (food to microorganisms ratio) showed the most consistent role (over the sludge retention time), values below 0.08 d-1 being associated with effluent PhC concentrations 26-45 % lower. In FNW, the nitrification role was statistically expressed by the alkalinity reduction, values above 40 % (i.e., Nt-removals >80 %) being associated with effluent PhC concentrations 23-61 % lower.

Twenty-four pharmaceutical active compounds (PhACs) were evaluated in the soft tissues of clams Ruditappes decussatus exposed along a 1.5-km dispersal gradient of the treated effluent from an urban wastewater treatment plant discharging in Ria Formosa, and compared with those in the marine waters and discharged effluents. The clams were exposed for 1 month, in June-July 2016, 2017 and 2018. PhACs were quantified by high performance liquid chromatography coupled to tandem mass spectrometry after the quick, easy, cheap, effective, rugged and safe (QuEChERS) method (clams) or solid-phase extraction (water samples). The most representative PhACs in the effluents and receiving waters (regardless of the tidal dilution effect) were diclofenac, carbamazepine and caffeine (on average <= 2 mu g/L) and only caffeine exhibited significant inter-annual differences, with higher values in 2017. In turn, the most bioaccumulated PhACs in clams were caffeine (0.54-27 ng/g wet weight, significantly higher in 2016) and acetaminophen (0.37-3.7 ng/g wet weight, significant lower in 2016). A multivariate principal component analysis showed (i) PhAC bioaccumulation primarily depended on biotic factors (clams length and weight), (ii) PhAC physicochemical properties Log Kow, pKa and water solubility interplaying with water abiotic variables were more relevant for explaining data variability in water than the physical dilution/tidal mixing, (iii) this process, reflected by the salinity gradient, had a tertiary role in data variation, responsible for spatial discrimination of marine waters. This study provides a better understanding of PhACs bioaccumulation by clams Ruditapes decussatus in real environmental conditions, under the influence of urban treated effluent dispersal in Ria Formosa coastal lagoon, a major producer of bivalves, ultimately disentangling key factors of PhAC bioaccumulation.

Although the evaluation of the uncertainty of an analytical method is a mandatory step in the method's validation, its applicability to the monitoring of trace compounds in complex samples is not simple, nor is it part of the routine of most laboratories, namely those dedicated to research. This manuscript focuses on the full validation of an analytical procedure for determining trace concentrations of twenty-four pharmaceutical active compounds (PhACs) in wastewaters using solid-phase extraction (SPE) and ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). The method optimization was performed on different wastewater matrices, namely influents and final effluents from two distinct wastewater treatment plants (WWTPs). Matrix effects and extraction efficiency (absolute recovery) of the developed method were determined. Validation was performed to obtain the method's linearity/working range, precision, trueness, method detection limits (MDLs) and method quantification limits (MQLs). The expanded uncertainty of the data obtained was estimated according to the requirements of international procedures dedicated to the expression of uncertainty. Different approaches for the estimation of uncertainty were applied. The validated method was used in the analysis of target PhACs in wastewater samples collected at two WWTPs. The obtained results facilitated the introduction of a validated method for routine measurement of PhACs in wastewater samples and allowed method accreditation by the competent national authority.

Powdered activated carbon (PAC) dosing to a conventional activated sludge (AS) reactor is a low-investment option for controlling pharmaceutical compounds (PhCs) in municipal wastewater treatment, but its advantages and limitations in real operating environments are not fully assessed. A 3-week PAC-AS full-scale test was conducted in an oxidation ditch plant to assess PAC impact on effluent quality (PhCs, dissolved organic carbon (DOC) and other parameters), energy consumption, sludge production and costs. DOC-normalized PAC doses of 0.7-2.6 mgPAC/mgDOC significantly reduced recalcitrant PhC discharge (e.g. by 63-83 % for carbamazepine and 67-69 % for diclofenac), the higher dose yielding a more reliable effluent quality. Effluent quality for total phosphorus, colour, organic matter and transmittance was also enhanced and no interference with nitrification, oxidation-reduction potential or dissolved oxygen in the oxidation ditch was observed, resulting in no energy consumption increase. PAC had no impact on effluent turbidity and mixed liquor suspended solids settleability, showing a positive effect on dewatered sludge dry weight and ultimately a 7-9 % increase on final sludge production. After stopping PAC dosing, PAC remaining in the return sludge presented adsorption capacity for some PhCs until it was completely out of the system. Estimated costs for PAC addition to AS-reactor compare favourably with literature data for PAC and GAC post-treatment and similarly with post-ozonation.

Some PhACs were not eliminated in WWTPs. Some PhACs were detected in upstream and downstream water samples. Some PhACs showed higher sensitivity for specific trophic levels.

The profile of pharmaceutical active compounds (PhACs) in two UWWTP was evaluated. The potential behaviour of PhACs on agriculture soils was evaluated based on their physicochemical properties and soil pH.

The use of biomarkers in bivalves has gained significance as a reliable method for the assessment of the presence and effect of contaminants in aquatic ecosystems. However, it is important to note that the biomarkers respond not only to contaminant loadings and bioavailability but also to environmental stress. Therefore, the association between biomarkers and contamination/pollution should be conducted cautiously as the environmental factors also affect their response. These factors should be integrated into the assessment of the response of the biomarkers. The potential impact of effluents from an urban wastewater treatment plant (UWWTP) on the Ruditapes decussatus clam specimens located 1.5 Km away in a surrounding area was evaluated. After one month of exposure, three biomarkers were analyzed, namely lipid peroxidation (LPO), acetylcholinesterase (AChE) and electron transport system (ETS). These parameters were also determined for a control group purchased from a local nursery, which had no influence from UWWTP, in order to compare the results obtained from both groups. The in situ physicochemical characterization of the exposure site (temperature, salinity, pH and dissolved oxygen) was evaluated together with nutritional parameters, morphometric measurements and condition index. The biochemical analysis showed that the activity levels of AChE fluctuated from 4.0 to 4.6 nmol/min g protein and that of LPO from 101.5 to 248.9 nmol MDA/g protein. Also, the ETS activity levels were in the range of 27.2 to 30.2 nmol O2/min g protein. The lipid peroxidation was found to be the most responsive biomarker toward the damage caused by environmental conditions on the clams.

ABSTRACT The occurrence of a representative range of pharmaceutical compounds (PhCs) in urban wastewaters was studied in two Portuguese water-scarce regions (Lisbon great area, and Faro – Algarve), involving 55 sampling campaigns during 2.5 years. The results show 16 PhCs (out of the 24 analysed) were quantified in all samples, with similar concentrations in both wastewaters. To understand to what extent seasonality affects PhC occurrence, a statistical analysis was conducted, and daily maximum air temperature was identified as a key factor, with a turning point of 20°C in Lisbon and 22°C in Faro. Some PhCs showed lower concentrations in colder (wet) months due to rainwater dilution (combined sewer systems), whereas others showed higher concentrations, reflecting an increased seasonal consumption and/or slower transformation under lower temperatures and/or shorter hydraulic retention times. Seasonal studies should therefore focus on temperature rather than on calendar seasons, increasingly uncertain due to climate change.

Pharmaceutical active compounds (PhACs) belonging to analgesics, antibiotics, and non-steroidal anti-inflammatory drugs (NSAIDs) therapeutic classes were monitored in wastewater influents and effluents from two Portuguese urban wastewater treatment plants (UWWTPs) for 24 months. Both facilities were chosen due to their effluents are discharged in highly touristic and sensitive areas, Tagus river and Ria Formosa coastal lagoon, respectively. Target PhACs, acetaminophen, diclofenac, ibuprofen, naproxen, sulfadiazine, and sulfamethoxazole were measured using solid-phase extraction (SPE) coupled to liquid chromatography tandem mass spectrometry (LC-MS/MS). PhACs occurrence in the influents was higher than 98%, with acetaminophen presenting the highest concentrations, with values between 16.3 µg/L and 124 µg/L. In the effluents, distinct behavior was observed with diclofenac and sulfamethoxazole showing recalcitrant characteristics, whereas acetaminophen, ibuprofen, and naproxen showed removal efficiencies above 95%. Acetaminophen and ibuprofen amount in influents showed consistently higher concentration levels in autumn (in Beirolas and Faro Nw UWWTPs) and winter (only in Beirolas UWWTP) seasons. These seasonal trends were observed to a greater extent in Beirolas UWWTP than in Faro Nw UWWTP. This study enabled the comprehensive definition of a behavior pattern for these target contaminants, contributing to better characterization and build-up a library of PhACs occurrence. It also allowed a robust seasonal profiling of the target compounds due to the high number of samples analyzed by each season and a longer monitoring campaign, making the obtained results more significant.

Presently, water quantity and quality problems persist both in developed and developing countries, and concerns have been raised about the presence of emerging pollutants (EPs) in water. The circular economy provides ways of achieving sustainable resource management that can be implemented in the water sector, such as the reuse of drinking water treatment sludges (WTSs). This study evaluated the potential of WTS containing a high concentration of activated carbon for the removal of two EPs: the steroid hormones 17β-estradiol (E2) and 17α-ethinylestradiol (EE2). To this end, WTSs from two Portuguese water treatment plants (WTPs) were characterised and tested for their hormone adsorbance potential. Both WTSs showed a promising adsorption potential for the two hormones studied due to their textural and chemical properties. For WTS1, the final concentration for both hormones was lower than the limit of quantification (LOQ). As for WTS2, the results for E2 removal were similar to WTS1, although for EE2, the removal efficiency was lower (around 50%). The overall results indicate that this method may lead to new ways of using this erstwhile residue as a possible adsorbent material for the removal of several EPs present in wastewaters or other matrixes, and as such contributing to the achievement of Sustainable Development Goals (SDG) targets.

Due to the high consumption and incorrect disposal of pharmaceutical active compounds (PhACs), they are recognized as contaminants of emerging concern. Wastewater treatment plants (WWTPs) may be inefficient in removing PhACs, therefore discharging them into surface waters. The removal efficiencies of five WWTPs located in the south of Portugal (Alentejo) were evaluated in 2020. Twenty-six PhACs were analyzed in wastewater influents, effluents, and surface waters, upstream and downstream of the WWTPs by solid-phase extraction (SPE) and ultra-performance liquid chromatography coupled with tandem mass detection (UPLC-MS/MS). The most representative PhACs in influents were acetaminophen, caffeine, naproxen, ibuprofen, and diclofenac with minimum-maximum concentrations of 49–225 µg/L, 26–46 µg/L, 5.9–13 µg/L, 5.2–22 µg/L, and 1.3–2.5 µg/L, respectively. For effluents, it was acetaminophen, caffeine, and diclofenac with minimum-maximum concentrations of 0.054–7.8 µg/L, 0.084–4.8 µg/L, and 0.28–3.3 µg/L, respectively. The highest removal efficiencies were observed for acetaminophen, sulfadiazine, cortisone, testosterone, metoprolol, and propranolol (100%). The lowest removal efficiencies were observed for carbamazepine (2.7%) and diclofenac (−13.2%). The risk quotient of sulfamethoxazole and diclofenac were higher than 1 for receiving waters, indicating they probably pose high risks to aquatic organisms.

To evaluate the potential impact of an urban wastewater-treatment plant on Ria Formosa coastal lagoon, a sentinel species, the clam Ruditapes decussatus, was exposed along a gradient of the effluent's dispersal for 1 mo. Three exposure sites were selected to study the responses of 3 biomarkers: electron transport system, acetylcholinesterase, and lipid peroxidation. As complementary data, morphometric measurements, condition index, and lipid and protein content were considered together with in situ physicochemical characterization of the sites (temperature, salinity, pH, and dissolved oxygen). Electron transport system activity levels were between 35.7 and 50.5 nmol O2 /min g protein, acetylcholinesterase activity levels ranged from 2.6 to 3.8 nmol/min g protein, and lipid peroxidation ranged from 174.7 to 246.4 nmol malondialdehyde/g protein. The exposure sites shaped the response not only of biomarkers but also of "health" parameters (protein, lipids, and condition index). Lipid peroxidation was the most responsive biomarker also associated with electron transport system, especially at the closest site to the urban wastewater-treatment plant. Because of the presence of complex mixtures of contaminants in urban effluents, biomarker responses can provide valuable information in environmental assessment. However, it is vital to identify all biological and ecological factors induced by the natural life cycle of clams. Abiotic factors can mask or overlap the response of biomarkers and should be considered in a multibiomarker approach. Environ Toxicol Chem 2021;40:272-283. © 2020 SETAC.

Data on different therapeutic classes of pharmaceutical compounds (PhCs) in clams or other bivalves living in natural conditions are scarce. The aim of this work was the optimization and validation of a method for the determination of PhCs in clams for further evaluation of any potential human exposure risk due to their consumption. A quick, easy, cheap, effective, rugged, and safe (QuEChERS) approach is proposed for sample clean-up and concentration of 24 PhCs in clams, with subsequent analysis by liquid chromatography-tandem mass spectrometry. This method showed a good linear range for all PhCs with determination coefficients (r2) between 0.9949 and 0.9993 and coefficients of variation (CVm) lower than 5.5%. This method allowed the quantification of target compounds at trace concentration levels (ng g−1), being the most abundant PhC in clam caffeine. This PhC was detected in more than 70% of samples with concentrations ranging from 0.10 to 12 ng g−1 wet weight.

The wisdom of the crowds effect (WoC) is a collective intelligence (CI) property by which, given a problem, a crowd is able to provide a solution better than that of any of its individuals. However, WoC is considered to require that participants are not priorly influenced by information received on the subject of the problem. Therefore it is important to have metrics that can identify the presence of influence in an experiment, so that who runs it can decide if the outcome is product of the WoC or of a cascade of individuals influencing others. In this paper we provide a set of metrics that can analyse a WoC experiment as a data stream and produce a clear indication of the presence of some influence. The results presented were obtained with real data from different information conditions, and are encouraging. The paper concludes with a discussion of relevant situations and points the most important steps that follow in this research.