Assessment of wastewater-borne pharmaceuticals in tissues and body fluids from riverine fish.

Riverine fish in densely populated areas is constantly exposed to wastewater-borne contaminants from effluent discharges. These can enter the organism through the skin, gills or by ingestion. Whereas most studies assessing the contaminant burden in exposed fish have focused either on muscle or a limited set of tissues. Here we set out to generate a more comprehensive overview of the distribution of pollutants across tissues by analyzing a panel of matrices including liver, kidney, skin, brain, muscle, heart, plasma and bile. To achieve a broad analyte coverage with a minimal bias towards a specific contaminant class, sample extracts from four fish species were analyzed by High-Performance Liquid Chromatography (HPLC) - high-resolution mass spectrometry (HRMS) for the presence of 600 wastewater-borne pharmaceutically active compounds (PhACs) with known environmental relevance in river water through a suspect-screening analysis. A total of 30 compounds were detected by suspect screening in at least one of the analyzed tissues with a clear prevalence of antidepressants. Of these, 15 were detected at confidence level 2.a (Schymanski scale), and 15 were detected at confidence level 1 following confirmation with authentic standards, which furthermore enabled their quantification. The detected PhACs confirmed with level 1 of confidence included acridone, acetaminophen, caffeine, clarithromycin, codeine, diazepam, diltiazem, fluoxetine, ketoprofen, loratadine, metoprolol, sertraline, sotalol, trimethoprim, and venlafaxine. Among these substances, sertraline stood out as it displayed the highest detection frequency. The values of tissue partition coefficients for sertraline in the liver, kidney, brain and muscle were correlated with its physicochemical properties. Based on inter-matrix comparison of detection frequencies, liver, kidney, skin and heart should be included in the biomonitoring studies of PhACs in riverine fish.

Analysis of pharmaceuticals in fish using ultrasound extraction and dispersive spe clean-up on que Z-Sep/C18 followed by LC-QToF-MS detection.

The presence of pharmaceutically active compounds (PhACs) in aquatic biota has been received much less attention than their presence in surface or waste water, and it was not until the mid-2000s, this gap started to be addressed. Here, we present SQUEEZe (Solid-liQuid Ultrasound Extraction with QuE Z-Sep/C18 as dispersive clean-up): a fast method for analysis of the trace 47 PhACs in fish muscle. Compared to our previously reported method [1], it offers alternatives with improvements in recoveries, number of analytes, sample volume and solvent used. The key aspects of this method are:•The ultrasound extraction was performed with acetonitrile/isopropanol 0.1% V/V formic acid. A clean-up step using QuE Z-Sep/C18 sorbents was employed to reduce lipid content of the extracts and further matrix effects in the detection of the analytes.•A HPLC separation with a Kinetex EVO C18 packed column in 11 min was optimized. MS and MS/MS data were collected using SWATH acquisition on the SCIEX X500R QTOF in (+)-ESI mode.•The method validated at 3 different concentrations levels: 5, 25 and 50 ng/g fish. It presented good intraday/interday reproducibility and absolute recoveries ≥ 60% for majority of analytes in composite homogenate muscle matrix of Squalius cephalus.•10 out 47 compounds were detected in fish samples.

Combining quantitative and qualitative approaches using Sequential Window Acquisition of All Theoretical Fragment-Ion methodology for the detection of pharmaceuticals and related compounds in river fish extracted using a sample miniaturized method.

A fast method for analysis of 47 pharmaceuticals active compounds (PhACs) in fish muscle has been developed and validated addressing the parameters accuracy, precision (intraday and interday), matrix effect at three spiking levels: 5, 25 and 50 ng PhAC g-1 in fish as well as linearity, limit of detection (LOD), limit of quantification (LOQ). Sixteen protocols were performed varying extraction techniques, solvents, sample filtration and clean-up step. The selected method was based on an ultrasound extraction with acidified mixture of acetonitrile and isopropyl alcohol followed by a clean-up step using Z-Sep/C18 sorbents. Quantitative analysis of the PhACs in the extracts was accomplished by UPLC- QTOF-MS using Sequential Window Acquisition of All Theoretical Fragment-Ion, SWATHTM acquisition technology. 90% Of the compounds presented extraction recoveries between 60 and 130% with LOQ between 0.2 and 11 ng g-1. The validated method was applied to the analysis of 32 muscle samples from thirteen different species of fish collected in four European river basins (Adige, Evrotas, Llobregat and Sava). A total of ten compounds were found in fish samples with diltiazem as the most frequently detected one followed by carbamazepine and caffeine. Additionally, by taking advantage of the information-rich mass spectral data from the SWATH mode acquisition, the raw data were reprocessed for the presence of the most prescribed 250 pharmaceuticals, metabolites, and drugs of abuse previously reported to occur in the aquatic environment. By considering the mass errors of the molecular ion (˂± 3 ppm) and one characteristic fragment ion (˂±10 ppm) as well as the Library score and the Formula Finder score of the data processing software six compounds were retrieved, and eventually four of them confirmed with authentic standards: cocaine and its metabolite benzoylecgonine, the stimulant nicotine, and the antibiotic ofloxacin. Two lipid regulators, lovastatin and simvastatin, were determined as a false positive.

Polluted water from an urban reservoir (Madín dam, México) induces toxicity and oxidative stress in Cyprinus carpio embryos.

The Madín Dam is a reservoir located in the municipalities of Naucalpan and Atizapán, in the metropolitan area adjacent to Mexico City. The reservoir supplies drinking water to nearby communities and provides an area for various recreational activities, including kayaking, sailing and carp fishing. Over time, the number of specimens of common carp has notably diminished in the reservoir, which receives direct domestic drainage from two towns as well as numerous neighborhoods along the Tlalnepantla River. Diverse studies have demonstrated that the pollutants in the water of the reservoir produce oxidative stress, genotoxicity and cytotoxicity in juvenile Cyprinus carpio, possibly explaining the reduction in the population of this species; however, it is necessary to assess whether these effects may also be occurring directly in the embryos. Hence, surface water samples were taken at five sites and pharmaceutical drugs, personal care products (especially sunscreens), organophosphate and organochlorine pesticides, and other persistent organic pollutants (e.g., polychlorinated biphenyls and polycyclic aromatic hydrocarbons) were identified. Embryos of C. carpio were exposed to the water samples to evaluate embryolethality, modifications in embryonic development, lipoperoxidation, the quantity of hydroperoxide and oxidized proteins, and antioxidant enzyme activity (superoxide dismutase, catalase and glutathione peroxidase). It was found that the polluted water of the Madín Dam gave rise to embryolethality, embryotoxicity, congenital abnormalities, and oxidative stress on the common carp embryos.

Development and validation of an analytical method for determination of pharmaceuticals in fish muscle based on QuEChERS extraction and SWATH acquisition using LC-QTOF-MS/MS system.

This study aimed at developing an analytical method for the extraction and quantification of 21 pharmaceutical actives compounds (PhACs) present in fish muscle. Using Norwegian Atlantic salmon as matrix, two extraction methods for PhACs were tested: ultrasound extraction (USE) using methanol (MeOH), acetonitrile (MeCN) or a mixture of MeCN:MeOH (1:1, v/v) as extracting solvents, and QuEChERS method using three different extraction salts. After selecting QuEChERS Original as extracting method of the analytes, three different clean-up methods were evaluated with respect to their efficiency to remove coextracted fat and lipids such as Enhanced Matrix Removal (EMR) and HLB prime. The dispersive-SPE EMR yielded the best recoveries for 21 of 27 analytes. PhACs were quantified by UPLC-MS/MS using SWATH acquisition mode. The method was validated in terms of recoveries, accuracy, linearity, precision, matrix effects at three levels of concentration: 25, 200 and 500 ng g-1 dw of fish muscle. For the majority of the analytes the recoveries were over 70%. Finally, the validated method was applied to natural riverine fish from the Evrotas river (Greece) and the Adige river (Italy) with positive findings for acetaminophen, propranolol, and venlafaxine reaching concentrations as high as 80 ng g-1 of muscle.

Removal of anti-inflammatory/analgesic pharmaceuticals from urban wastewater in a pilot-scale A2O system: Linking performance and microbial population dynamics to operating variables.

In this study, the removal rates of eight anti-inflammatory and/or analgesic pharmaceuticals, AIAPs (acetaminophen, ibuprofen, naproxen, ketoprofen, diclofenac, codeine, indomethacin and propyphenazone) were assessed in a pilot-scale A2O system (including anaerobic/anoxic/aerobic zones), long term operated during two experimental phases using different sets of environmental conditions and operating parameters. qPCR was used to quantify the absolute abundances of total Bacteria, total Archaea, mycolic-acid containing filamentous Actinobacteria (Mycolata) and Fungi within the activated sludge microbial community developed in the system. Multivariate analyses and Spearman correlation coefficients were used in search of significant links among the removal rates of the AIAPs, the abundances of the targeted microbial groups in the activated sludge, and the changes of environmental/operating variables in the A2O system. Improved removal efficiencies of several of the AIAPs analyzed (acetaminophen, ibuprofen, naproxen, ketoprofen) were correlated to higher organic load in the influent water, higher concentration of mixed liquor suspended solids (MLSS), lower temperature and lower food-to-microorganisms ratio (F/M). Removal efficiencies of several pharmaceuticals correlated with increased abundances of Mycolata in the A2O system, pointing at this group of bacteria as candidate key players for AIAPs removal in activated sludge.