Consumption of illicit drugs and benzodiazepines in six Spanish cities during different periods of the COVID-19 pandemic.

Wastewater-based epidemiology (WBE) can provide objective and real time information about the use of addictive substances. A national study was conducted by measuring the most consumed illicit drugs, other drugs whose consumption is not so widespread but has increased significantly in recent years, and benzodiazepines in untreated wastewater from seven wastewater treatment plants (WWTPs) in six Spanish cities. Raw composite wastewater samples were collected from December 2020 to December 2021, a period in which the Spanish and regional governments adopted different restriction measures to contain the spread of the COVID-19 pandemic. Samples were analyzed using a validated analytical methodology for the simultaneous determination of 18 substances, based on solid-phase extraction and liquid-chromatography tandem mass spectrometry. Except for heroin, fentanyl, 6-acetylmorphine and alprazolam, all the compounds were found in at least one city and 9 out of 18 compounds were found in all the samples. In general, the consumption of illicit drugs was particularly high in one of the cities monitored in December 2020, when the restrictions were more severe, especially for cannabis and cocaine with values up to 46 and 6.9 g/day/1000 inhabitants (g/day/1000 inh), respectively. The consumption of MDMA, methamphetamine and mephedrone was notably higher in June 2021, after the end of the state of alarm, in the biggest population investigated in this study. Regarding the use of benzodiazepines, the highest mass loads corresponded to lorazepam. This study demonstrates that WBE is suitable for complementing epidemiological studies about the prevalence of illicit drugs and benzodiazepines during the COVID-19 pandemic restrictions.

Miniaturized method for the quantification of persistent organic pollutants and their metabolites in HepG2 cells: assessment of their biotransformation.

Biotransformation can greatly influence the accumulation and, subsequently, toxicity of substances in living beings. Although traditionally these studies to quantify metabolization of a compound have been carried out with in vivo species, currently, in vitro test methods with very different cell lines are being developed for their evaluation. However, this is still a very limited field due to multiple variables of a very diverse nature. So, an increasing number of analytical chemists are working with cells or other similar biological samples of very small size. This makes it necessary to address the development of analytical methods that allow determining their concentration both inside the cells and in their exposure medium. The aim of this study is to develop a set of analytical methodologies for the quantification of polycyclic aromatic hydrocarbons, PAHs (phenanthrene, PHE), and polybrominated diphenyl ethers, PBDEs (2,2',4,4'-tetrabromodiphenyl ether, BDE-47), and their major metabolites in cells and their exposure medium. Analytical methodologies, based on miniaturized ultrasound probe-assisted extraction, gas chromatography-mass spectrometry-microelectron capture detector (GC-MS-µECD), and liquid chromatography-fluorescence detector (LC-FL) determination techniques, have been optimized and then applied to a biotransformation study in HepG2 at 48 h of exposure. Significant concentrations of the major metabolites of PHE (1-OH, 2-OH, 3-OH, 4-OH-, and 9-OH-PHE) and BDE-47 (5-MeO-, 5-OH-, and 3-OH-BDE-47) were detected and quantified inside the cells and in the exposure medium. These results provide a new method for determination and improve information on the metabolization ratios for a better knowledge of the metabolic pathways and their toxicity.

Bioaccumulation and Biotransformation of BDE-47 Using Zebrafish Eleutheroembryos (Danio rerio).

Polybrominated diphenyl ethers (PBDEs) are well-known endocrine disrupting chemicals identified as organic persistent pollutants. Their metabolites OH-BDE and MeO-BDE have been reported to be potentially more toxic than the postulated precursor PBDEs. One of the most predominant congeners of PBDEs in the environment is BDE-47, due to its high presence in industrially used mixtures. In the present study, the bioaccumulation and biotransformation of BDE-47 into its major metabolites is evaluated using zebrafish (Danio rerio) eleutheroembryos adapting a previously developed alternative method to bioconcentration official guideline Organisation for Economic Co-ordination and Development 305, which reduces the animal suffering, time, and cost. For the simultaneous determination of BDE-47 and its metabolites in larvae and exposure medium, and considering the polarity difference of the analytes and the small sample size, the development of a validated analytical method is a step to ensure quality results. In the present study, an ultrasound-assisted extraction followed by a solid phase extraction dispersive clean-up step and gas chromatography-mass spectrometry-microelectron capture detector (GC-MS-µECD) with a previous derivatization process was optimized and validated. Bioconcentration factors (BCFs) were calculated using a first-order one-compartment toxicokinetic model. The profiles found show rapid absorption in the first hours of larval development and great bioaccumulative capacity, finding BCFs of 7294 ± 899 and 36 363 ± 5702 at nominal concentrations of 10 and 1 µg L-1 , respectively. Metabolization studies show increasing concentrations of the metabolites BDE-28, 2'-OH-BDE-28, and 5-MeO-BDE-47 throughout the exposure time. The results obtained show the feasibility of the method for bioaccumulation and open up the possibility of metabolic studies with zebrafish eleutheroembryos, which is a very underdeveloped field without official testing or regulation. Environ Toxicol Chem 2023;42:835-845. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Development of a method for assessing the accumulation and metabolization of antidepressant drugs in zebrafish (Danio rerio) eleutheroembryos.

Antidepressant drugs are widely used for the treatment of common mental or other psychiatric disorders such as depression, which affect about 121 million people worldwide. This widespread use has contributed to the input of these pharmaceuticals and their metabolites into the environment. The aim of this work was to develop an analytical method to quantify the most widely used antidepressant drugs, selective serotonin reuptake inhibitors (SSRI), and their main metabolites in the environment. For this, a new and reliable miniaturized extraction method based on dispersive SPE cleanup procedure for extraction of SSRI followed by derivatization with n-heptafluorobutyrylimidazole, and detection by GC-MS was developed. The methodology, including a first-order one-compartment model, was then applied to a bioconcentration study in zebrafish (Danio rerio) eleutheroembryos. The results showed low bioaccumulation of these compounds; however, a biotransformation evidence of the parent compounds into their metabolites was observed after 6 h of exposure. These results indicate the need to integrate metabolic transformation rates to fully model and understand the bioaccumulation patterns of SSRI and their metabolites. Graphical abstract.

In vivo quantification of volatile organoselenium compounds released by bacteria exposed to selenium with HS-SPME-GC-MS. Effect of selenite and selenium nanoparticles.

Quantification of volatile organoselenium species released by Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), after their growth in the presence of 1 and 2 mg Se·L-1 as both selenite and chitosan-modified selenium nanoparticles (Ch-SeNPs), was achieved by the application of a method based on headspace solid-phase microextraction (HS-SPME) and in-fiber internal standardization, combined with gas chromatography coupled to mass spectrometry (GC-MS). This method consisted of an initial extraction of the released volatile organoselenium compounds on the SPME fiber, followed by the extraction of internal standard (IS), deuterated dimethyl sulfide (d6-DMS), on the same fiber before its desorption at the injection port of GC-MS. The results showed that the biotransformation of selenite and Ch-SeNPs into volatile organoselenium compounds was dependent on both the type of bacterial species and the chemical form of selenium (Se) administered. In this sense, E. coli was able to biotransform both selenite and Ch-SeNPs into dimethylselenium (DMSe) and dimethyldiselenium (DMDSe) while S. aureus, biotransformed selenite into DMSe and DMDSe and, Ch-SeNPs only into DMDSe. Additionally, the formation of a volatile mixed sulfur/selenium compound, dimethyl selenenyl sulfide (DMSeS), from Se in nanoparticulated form has been detected for the first time.

Insights into the accumulation and transformation of Ch-SeNPs by Raphanus sativus and Brassica juncea: Effect on essential elements uptake.

Selenium (Se) at very low doses has important functions for humans. Unfortunately, the low levels of Se in soils in various regions of the world have implemented the agronomic biofortification of crops by applying Se-enriched fertilizers (mainly based on selenate). Lately, the use of nanofertilizers is growing in interest as their low size reduces the amount of chemicals and minimizes nutrient losses in comparison with conventional bulk fertilizers. However, the knowledge on their fate and environmental impact is still scarce. This study aims to evaluate the biotransformation of chitosan-modified Se nanoparticles (Ch-SeNPs) as well as their effect on the metabolism of essential metals (Fe, Cu, Zn and Mo) when applied to hydroponic cultivation of R. sativus and B. juncea. In house-synthesized Ch-SeNPs were characterized in both synthesis and hydroponic culture media by transmission electron microscopy (TEM), dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA). The composition of one-tenth strength Hoagland's solution did not affect the size, shape and concentration in number of particles per mL of Ch-SeNPs. The plants were grown inside a box at 25 °C during the months of May-July in 2018. After a week of treatment with Ch-SeNPs, plants were harvested and divided into roots and aerial part. The biotransformation of Ch-SeNPs was evaluated through a process of enzymatic hydrolysis and subsequent analysis by HPLC-ICP-MS and HPLC-ESI-MS/MS. The results confirmed the transformation of Ch-SeNPs to seleno-amino acids: Selenomethionine (SeMet), Semethylselenocysteine (SeMetSeCys) and É£-glutamyl-Se-MetSeCys. Moreover, Multiple-way analysis of variance (ANOVA) and principal component analysis (PCA) showed that, regardless the plant species, Ch-SeNPs supplementation affected the absorption of Zn.

In-vivo solid phase microextraction for quantitative analysis of volatile organoselenium compounds in plants.

A new calibration method based on the use of headspace solid-phase microextraction (HS-SPME) and in-fiber internal standardization, combined with gas chromatography coupled to mass spectrometry (GC/MS) was developed for quantifying Se volatile organic species released by plants exposed to chitosan-modified selenium nanoparticles (Cs-SeNPs). The effect of several parameters affecting extraction and separation of the selected organic species of selenium (dimethylselenium (DMSe), diethylselenium (DESe) and dimethyldiselenium (DMDSe)) and deuterated dimethyl sulphide (d6-DMS) employed as internal standard were studied and optimized using an experimental design. The developed methodology was applied for quantifying the volatile selenium compounds produced over time by the plant species Raphanus sativus and Brassica juncea grown in hydroponic solution containing 5 mg Se L-1 in the form Cs-SeNPs. The procedure employed consisted in two steps. Volatile selenium species released from the plants were first extracted in the SPME fiber located at the headspace of a box with a fixed volume. Subsequently, the internal standard placed in a vial subjected to the same conditions as plants was extracted on the same fiber than the one previously used for extracting selenium compounds. Finally the extracted compounds were separated and analyzed by GC/MS. Results evidenced Cs-SeNPs biotransformation into DMSe and DMDSe by both plants species during growing stage, in amounts of the order of ng. Additionally, the resulting data were submitted to multifactorial ANOVA to evaluate the influence of plant type and time of exposure to Cs-SeNPs on the production of volatile selenium compounds.

Impact of selenium co-administration on methylmercury exposed eleutheroembryos and adult zebrafish (Danio rerio): Changes in bioaccumulation and gene expression.

Mercury still represents one of the most hazardous threats for the aquatic ecosystem due to its high toxicity, and the fact that it can be easily incorporated into the food chain by accumulation in fish as MeHg. On the other hand, selenium is a micronutrient that is part of different antioxidant enzymes that regulate the cellular redox state, and whose complex interaction with Hg has been extensively studied from a toxicological point of view. In order to evaluate the protective effect of Se(IV) co-administration against MeHg accumulation and toxicity, we have selected an in-vivo model at two developmental stages: zebrafish eleutheroembryos and adult fish. Embryos were exposed during 48 h to MeHg (5 or 25 µg/l) and a concentration of Se (IV) representing a molar ratio close to one (2.5 or 12.5 µg/l), while adult zebrafish were exposed during 72 h to either 25 µg/l of MeHg alone or co-exposed with 12.5 µg/l of Se (IV). A significant decrease in MeHg bioaccumulation factor was observed in eleutheroembryos co-exposed to Se(IV). A time-dependent accumulation of MeHg was observed in all the analyzed organs and tissues of adult fish, which was significantly reduced in the muscular tissue and the intestine by Se(IV) co-administration. However, such protection against MeHg bioaccumulation was not maintained in the brain and liver. The data derived from the gene expression analysis also demonstrated the protective effect of Se(IV) against MeHg-induced oxidative stress and the activation of different defense mechanisms by Se(IV) co-administration.

Detection of exposure effects of mixtures of heavy polycyclic aromatic hydrocarbons in zebrafish embryos.

In this study we evaluated the exposure effects of mixtures of five polycyclic aromatic hydrocarbons (PAHs); namely, benzo[a]anthracene, benzo[a]pyrene, benzo[b]fluoranthene, benzo[k]fluoranthene and chrysene on zebrafish embryos. Supplementation of the exposure media with 0.45% dimethyl sulfoxide and 50 ppm of Tween 20 could guarantee the solubilization and stabilization of the PAHs up to 24 h without affecting the embryos development. The exposure effects were tested by detecting the differential expression of a number of genes related to the aryl hydrocarbon receptor gene battery. Effects were detectable already after 6 h of exposure. After 24 h of exposure, all PAHs, except for benzo[a]anthracene, acted as potent inducers of the gene cyp1a1. Benzo[k]fluoranthene was the major inducer; the effect caused by the mixture at the lower concentration tested (1 ng ml-1 ) was dominated by its presence. However, in the mixture at the highest concentration tested (10 ng ml-1 ) it caused less induction and was not dominant. No significant bioaccumulation values were detected on embryos exposed to the PAHs tested in this study; however, the results obtained, indicated that PAHs undergo a very rapid metabolization inside the embryos, and that those biotransformation products yield changes on the expression of genes involved in the aryl hydrocarbon receptor pathway. Future work should focus on identification of the PAH metabolization products and on the effect of these metabolites on toxicity. Copyright © 2016 John Wiley & Sons, Ltd.

Evaluation of chromatographic columns packed with semi- and fully porous particles for benzimidazoles separation.

The behavior of 15 benzimidazoles, including their main metabolites, using several C18 columns with standard or narrow-bore diameters and different particle size and type were evaluated. These commercial columns were selected because their differences could affect separation of benzimidazoles, and so they can be used as alternative columns. A simple screening method for the analysis of benzimidazole residues and their main metabolites was developed. First, the separation of benzimidazoles was optimized using a Kinetex C18 column; later, analytical performances of other columns using the above optimized conditions were compared and then individually re-optimized. Critical pairs resolution, analysis run time, column type and characteristics, and selectivity were considered for chromatographic columns comparison. Kinetex XB was selected because it provides the shortest analysis time and the best resolution of critical pairs. Using this column, the separation conditions were re-optimized using a factorial design. Separations obtained with the different columns tested can be applied to the analysis of specific benzimidazoles residues or other applications.

Bioaccumulation of ionic titanium and titanium dioxide nanoparticles in zebrafish eleutheroembryos.

The production of titanium dioxide nanoparticles (TiO(2) NPs) for commercial applications has greatly increased over the last years and consequently the potential risk for human health. There is a growing awareness of the need to understand the behavior and influence these nanoparticles exert on the environment. Bioaccumulation serves as a good integrator to assess chemical exposure in aquatic systems and is dependent on factors, such as the exposure routes, diet and the aqueous medium. We analyzed the experimental bioaccumulation capability of ionic titanium and TiO(2) NPs by zebrafish (Danio rerio) eleutheroembryos through bioconcentration factors (BCFs), after 48 or 72 h of exposure. The stability of both chemical forms in an aquatic medium was fully characterized for further bioaccumulation studies. Several stabilizing agents (humic acids, soluble starch, polyethylene glycol, Na(4)P(2)O(7) and Na(2)HPO(4)) for anatase and rutile, the two allotrophs of TiO(2) NPs, were evaluated to check the evolution of the aggregation process. Around 60% of TiO(2) NPs remained disaggregated under simulated environmental conditions with the addition of 50 mg L(-1) of humic acids. However, the presence of eleutheroembryos in the exposure medium increased TiO(2) NPs aggregation in the experimental tests. The BCFs values obtained in all cases were <100, which classifies ionic titanium and TiO(2) NPs as non-bioaccumulative substances, under the REACH regulations.