Detection of Bromochloro Alkanes in Indoor Dust Using a Novel CP-Seeker Data Integration Tool.

Bromochloro alkanes (BCAs) have been manufactured for use as flame retardants for decades, and preliminary environmental risk screening suggests they are likely to behave similarly to polychlorinated alkanes (PCAs), subclasses of which are restricted as Stockholm Convention Persistent Organic Pollutants (POPs). BCAs have rarely been studied in the environment, although some evidence suggests they may migrate from treated-consumer materials into indoor dust, resulting in human exposure via inadvertent ingestion. In this study, BCA-C14 mixture standards were synthesized and used to validate an analytical method. This method relies on chloride-enhanced liquid chromatography-electrospray ionization-Orbitrap-high resolution mass spectrometry (LC-ESI-Orbitrap-HRMS) and a novel CP-Seeker integration software package for homologue detection and integration. Dust sample preparation via ultrasonic extraction, acidified silica cleanup, and fractionation on neutral silica cartridges was found to be suitable for BCAs, with absolute recovery of individual homologues averaging 66 to 78% and coefficients of variation ≤10% in replicated spiking experiments (n = 3). In addition, a total of 59 indoor dust samples from six countries, including Australia (n = 10), Belgium (n = 10), Colombia (n = 10), Japan (n = 10), Thailand (n = 10), and the United States of America (n = 9), were analyzed for BCAs. BCAs were detected in seven samples from the U.S.A., with carbon chain lengths of C8, C10, C12, C14, C16, C18, C24 to C28, C30 and C31 observed overall, though not detected in samples from any other countries. Bromine numbers of detected homologues in the indoor dust samples ranged Br1-4 as well as Br7, while chlorine numbers ranged Cl2-11. BCA-C18 was the most frequently detected, observed in each of the U.S.A. samples, while the most prevalent degrees of halogenation were homologues of Br2 and Cl4-5. Broad estimations of BCA concentrations in the dust samples indicated that levels may approach those of other flame retardants in at least some instances. These findings suggest that development of quantification strategies and further investigation of environmental occurrence and health implications are needed.

Trapped and drift-tube ion-mobility spectrometry for the analysis of environmental contaminants: Comparability of collision cross-section values and resolving power.

RATIONALE: Ion-mobility (IM)-derived collision cross-section (CCS) values can serve as a valuable additional identification parameter within suspect and non-target screening studies of environmental contaminants. However, these applications require to assess the reproducibility of CCS calculations between different IM set-ups. Especially for the comparison of trapped and drift-tube IM (TIMS/DTIM) derived CCS values, data for environmental applications is lacking. METHODS: The presented study assessed the bias of TIMS derived CCSN2 (TIMSCCSN2) values of 48 environmental contaminants from three classes in comparison to a previously established DTIM database. Based on two sets of isomeric bisphenols, the resolving power of both systems was compared, addressing the instrumental settings which influence the resolution of TIMS measurements. RESULTS: For 91% of the datapoints, bias between TIMSCCSN2 and DTCCSN2 values (latter set as reference) were < 2%, indicating a good inter-platform reproducibility. TIMS resolving power was dependent on the selected mobility window and ramping times whereby a resolution of up to 116 was achieved. Similar resolving power was observed for multiplexed DTIMS data if a high-resolution post-processing step was implemented. CONCLUSIONS: These results provide valuable insights in CCSN2 reproducibility facilitating database transfer in future TIMS based studies. Knowledge on the influence of acquisition settings on robustness of TIMSCCSN2 calculations and resolving power can ease method development supporting efficient development and reliable identifications of emerging environmental contaminants.

In Vitro Metabolism of Quaternary Ammonium Compounds and Confirmation in Human Urine by Liquid Chromatography Ion-Mobility High-Resolution Mass Spectrometry.

Quaternary ammonium compounds (QACs) are high-production chemicals used as cleaning and disinfecting agents. Due to their ubiquitous presence in the environment and several toxic effects described, human exposure to these chemicals gained increasing attention in recent years. However, very limited data on the biotransformation of QACs is available, hampering exposure assessment. In this study, three QACs (dimethyl dodecyl ammonium, C10-DDAC; benzyldimethyl dodecylammonium, C12-BAC; cetyltrimethylammonium, C16-ATMAC) commonly detected in indoor microenvironments were incubated with human liver microsomes and cytosol (HLM/HLC) simulating Phase I and II metabolism. Thirty-one Phase I metabolites were annotated originating from 19 biotransformation reactions. Four metabolites of C10-DDAC were described for the first time. A detailed assessment of experimental fragmentation spectra allowed to characterize potential oxidation sites. For each annotated metabolite, drift-tube ion-mobility derived collision cross section (DTCCSN2) values were reported, serving as an additional identification parameter and allowing the characterization of changes in DTCCSN2 values following metabolism. Lastly, eight metabolites, including four metabolites of both C12-BAC and C10-DDAC, were confirmed in human urine samples showing high oxidation states through introduction of up to four oxygen atoms. This is the first report of higher oxidized C10-DDAC metabolites in human urine facilitating future biomonitoring studies on QACs.

Occurrence of short- and ultra-short chain PFAS in drinking water from Flanders (Belgium) and implications for human exposure.

SHORT: and ultra-short chain perfluoroalkyl substances (S- and US-PFAS) are alternatives for the long-chain PFAS which have been more regulated over time. They are highly mobile in the environment and can easily reach drinking water sources which can become an important human exposure route. Furthermore, there have been growing concerns about the presence of PFAS in Flanders. Because of this, human exposure to S- and US-PFAS through Flemish drinking water was evaluated in this study. For this purpose, the presence of 2 S-PFAS (PFBS and PFBA) and 5 US-PFAS (PFPrS, PFEtS, TFMS, PFPrA and TFA) was investigated in 47 tap water samples, collected from different Flemish provinces, and 16 bottled waters purchased in Flanders. Out of the 7 target PFAS, 4 (PFBA, PFBS, PFPrS and PFEtS) were detected at concentrations above LOQ in tap water. In bottled water, only TFMS was present above its LOQ. PFAS concentrations in all analyzed water samples ranged from <0.7 to 7.3 ng/L for PFBS, <0.03-15.0 ng/L for TFMS and <0.9-12.0 ng/L for PFBA. PFPrS was only detected once above its LOQ, at 0.6 ng/L. No value could be reported for PFPrA due to high procedural blanks resulting in a high LOQ, nor for TFA due to high matrix effect. No significant differences in PFAS concentrations were seen in tap water among different drinking water companies, provinces, nor between the two types of analyzed bottled water (natural mineral water vs spring water). The use of a commercial carbon filter significantly reduced the median concentrations of the studied compounds in tap water. Finally, it was estimated that the presence of S- and US-PFAS in Flemish drinking water does not pose an immediate threat to human health, as concentrations were at least two orders of magnitude below the available guidance values.

Exploring the hidden chemical landscape: Non-target and suspect screening analysis for investigating solid waste-associated environments.

Solid waste is an inevitable consequence of urbanization. It can be safely managed in municipal landfills and processing plants for volume reduction or material reuse, including organic solid waste. However, solid waste can also be discarded in (un-)authorized dumping sites or inadvertently released into the environment. Legacy and emerging contaminants have the potential to leach from solid waste, making it a significant pathway to the environment. Non-target screening (NTS) and suspect screening analysis (SSA) have become helpful tools in environmental science for the simultaneous analysis of a wide range of chemical compounds. However, the application of these analytical approaches to environmental samples related to Raw or Processed Solid Waste (RPSW) has been largely neglected so far. This perspective review examines the potential and policy relevance of NTS and SSA applied to waste-related samples (liquid, gaseous and solid). It addresses the hurdles associated with the chemical safety of solid waste accumulation, processing, and reuse, and the need for landfill traceability, as well as effectiveness of leachate treatments. We reviewed the current applications of NTS and SSA to environmental samples of RPSW, as well as the potential adaptation of NTS and SSA techniques from related fields, such as oilfield and metabolomics, to the solid waste domain. Despite the ongoing technical challenges, this review highlights the significant potential for the implementation of NTS and SSA approaches in solid waste management and related scientific fields and provides support and guidance to the regulatory authorities.

HBM4EU E-waste study: Assessing persistent organic pollutants in blood, silicone wristbands, and settled dust among E-waste recycling workers in Europe.

E-waste recycling is an increasingly important activity that contributes to reducing the burden of end-of-life electronic and electrical apparatus and allows for the EU's transition to a circular economy. This study investigated the exposure levels of selected persistent organic pollutants (POPs) in workers from e-waste recycling facilities across Europe. The concentrations of seven polychlorinated biphenyls (PCBs) and eight polybrominated diphenyl ethers (PBDEs) congeners were measured by GC-MS. Workers were categorized into five groups based on the type of e-waste handled and two control groups. Generalized linear models were used to assess the determinants of exposure levels among workers. POPs levels were also assessed in dust and silicone wristbands (SWB) and compared with serum. Four PCB congeners (CB 118, 138, 153, and 180) were frequently detected in serum regardless of worker's category. With the exception of CB 118, all tested PCBs were significantly higher in workers compared to the control group. Controls working in the same company as occupationally exposed (Within control group), also displayed higher levels of serum CB 180 than non-industrial controls with no known exposures to these chemicals (Outwith controls) (p < 0.05). BDE 209 was the most prevalent POP in settled dust (16 µg/g) and SWB (220 ng/WB). Spearman correlation revealed moderate to strong positive correlations between SWB and dust. Increased age and the number of years smoked cigarettes were key determinants for workers exposure. Estimated daily intake through dust ingestion revealed that ΣPCB was higher for both the 50th (0.03 ng/kg bw/day) and 95th (0.09 ng/kg bw/day) percentile exposure scenarios compared to values reported for the general population. This study is one of the first to address the occupational exposure to PCBs and PBDEs in Europe among e-waste workers through biomonitoring combined with analysis of settled dust and SWB. Our findings suggest that e-waste workers may face elevated PCB exposure and that appropriate exposure assessments are needed to establish effective mitigation strategies.

Logistic Regression Analysis of LC-MS/MS Data of Monomers Eluted from Aged Dental Composites: A Supervised Machine-Learning Approach.

Compound identification by database searching that matches experimental with library mass spectra is commonly used in mass spectrometric (MS) data analysis. Vendor software often outputs scores that represent the quality of each spectral match for the identified compounds. However, software-generated identification results can differ drastically depending on the initial search parameters. Machine learning is applied here to provide a statistical evaluation of software-generated compound identification results from experimental tandem MS data. This task was accomplished using the logistic regression algorithm to assign an identification probability value to each identified compound. Logistic regression is usually used for classification, but here it is used to generate identification probabilities without setting a threshold for classification. Liquid chromatography coupled with quadrupole-time-of-flight tandem MS was used to analyze the organic monomers leached from resin-based dental composites in a simulated oral environment. The collected tandem MS data were processed with vendor software, followed by statistical evaluation of these results using logistic regression. The assigned identification probability to each compound provides more confidence in identification beyond solely by database matching. A total of 21 distinct monomers were identified among all samples, including five intact monomers and chemical degradation products of bisphenol A glycidyl methacrylate (BisGMA), oligomers of bisphenol-A ethoxylate methacrylate (BisEMA), triethylene glycol dimethacrylate (TEGDMA), and urethane dimethacrylate (UDMA). The logistic regression model can be used to evaluate any database-matched liquid chromatography-tandem MS result by training a new model using analytical standards of compounds present in a chosen database and then generating identification probabilities for candidates from unknown data using the new model.

Investigating the Potential of Drift Tube Ion Mobility for the Analysis of Oxidized Lipids.

Epilipids, a subset of the lipidome that comprises oxidized, nitrated, and halogenated lipid species, show important biochemical activity in the regulation of redox lipid metabolism by influencing cell fate decisions, including death, health, and aging. Due to the large chemical diversity, reversed-phase liquid chromatography-high-resolution mass spectrometry (RPLC-HRMS) methods have only a limited ability to separate numerous isobaric and isomeric epilipids. Ion mobility spectrometry (IMS) is a gas-phase separation technique that can be combined with LC-HRMS to improve the overall peak capacity of the analytical platform. Here, we illustrate the advantages and discuss the current limitations of implementing IMS in LC-HRMS workflows for the analysis of oxylipins and oxidized complex lipids. Using isomeric mixtures of oxylipins, we demonstrated that while deprotonated ions of eicosanoids were poorly resolved by IMS, sodium acetate and metal adducts (e.g., Li, Na, Ag, Ba, K) of structural isomers often showed ΔCCS% above 1.4% and base peak separation with high-resolution demultiplexing (HRDm). The knowledge of the IM migration order was also used as a proof of concept to help in the annotation of oxidized complex lipids using HRDm and all-ion fragmentation spectra. Additionally, we used a mixture of deuterium-labeled lipids for a routine system suitability test with the purpose of improving harmonization and interoperability of IMS data sets in (epi)lipidomics.

Exposure to a Mixture of Endocrine-Disrupting Chemicals and Metabolic Outcomes in Belgian Adolescents.

Childhood exposure to endocrine-disrupting chemicals (EDCs), either alone or in mixtures, may affect metabolic outcomes, yet existing evidence remains inconclusive. In our study of 372 adolescents from the Flemish Environment and Health Study (FLEHS IV, 2017-2018), we measured 40 known and suspected EDCs and assessed metabolic outcomes, including body mass index z-score (zBMI), abdominal obesity (AO), total cholesterol (TC), and triglycerides (TG). We applied Bayesian kernel machine regression (BKMR) and Bayesian penalized horseshoe regression for variable selection and then built multivariate generalized propensity score (mvGPS) models to provide an overview of the effects of selected EDCs on metabolic outcomes. As a result, BKMR and horseshoe together identified five EDCs associated with zBMI, three with AO, three with TC, and five with TG. Through mvGPS analysis, monoiso-butyl phthalate (MIBP), polychlorinated biphenyl (PCB-170), and hexachlorobenzene (HCB) each showed an inverse association with zBMI, as did PCB-170 with AO. Copper (Cu) was associated with higher TC and TG, except in boys where it was linked to lower TG. Additionally, monoethyl phthalate (MEP) and monobenzyl phthalate (MBzP) were associated with higher TG. To conclude, our findings support the association between certain chemicals (Cu, MEP, and MBzP) and elevated lipid levels, aligning with prior studies. Further investigation is needed for sex-specific effects.

Association of Endocrine-Disrupting Chemicals with All-Cause and Cause-Specific Mortality in the U.S.: A Prospective Cohort Study.

Wide exposure to endocrine-disrupting chemicals (EDCs) poses a great risk on human health. However, few large-scale cohort studies have comprehensively estimated the association between EDCs exposure and mortality risk. This study aimed to investigate the association of urinary EDCs exposure with mortality risk and quantify attributable mortality and economic loss. Multivariable Cox proportional hazards regression models were performed to investigate the association of 38 representative EDCs exposure with mortality risk in the National Health and Nutrition Examination Survey (NHANES). During a median follow-up of 7.7 years, 47,279 individuals were enrolled. All-cause mortality was positively associated with 1-hydroxynaphthalene, 2-hydroxynaphthalene, cadmium, antimony, cobalt, and monobenzyl phthalate. Cancer mortality was positively associated with cadmium. Cardiovascular disease (CVD) mortality was positively associated with 1-hydroxynaphthalene, 2-hydroxynaphthalene, and 2-hydroxyfluorene. Nonlinear U-shaped relationships were found between all-cause mortality and cadmium and cobalt, which was also identified between 2-hydroxyfluorene and CVD mortality. J-shaped association of cadmium exposure with cancer mortality was also determined. EDCs exposure may cause 56.52% of total deaths (1,528,500 deaths) and around 1,897 billion USD in economic costs. Exposure to certain phthalates, polycyclic aromatic hydrocarbons, phytoestrogens, or toxic metals, even at substantially low levels, is significantly associated with mortality and induces high economic costs.

Untargeted hair lipidomics: comprehensive evaluation of the hair-specific lipid signature and considerations for retrospective analysis.

Lipidomics investigates the composition and function of lipids, typically employing blood or tissue samples as the primary study matrices. Hair has recently emerged as a potential complementary sample type to identify biomarkers in early disease stages and retrospectively document an individual's metabolic status due to its long detection window of up to several months prior to the time of sampling. However, the limited coverage of lipid profiling presented in previous studies has hindered its exploitation. This study aimed to evaluate the lipid coverage of hair using an untargeted liquid chromatography-high-resolution mass spectrometry lipidomics platform. Two distinct three-step exhaustive extraction experiments were performed using a hair metabolomics one-phase extraction technique that has been recently optimized, and the two-phase Folch extraction method which is recognized as the gold standard for lipid extraction in biological matrices. The applied lipidomics workflow improved hair lipid coverage, as only 99 species could be annotated using the one-phase extraction method, while 297 lipid species across six categories were annotated with the Folch method. Several lipids in hair were reported for the first time, including N-acyl amino acids, diradylglycerols, and coenzyme Q10. The study suggests that hair lipids are not solely derived from de novo synthesis in hair, but are also incorporated from sebum and blood, making hair a valuable matrix for clinical, forensic, and dermatological research. The improved understanding of the lipid composition and analytical considerations for retrospective analysis offers valuable insights to contextualize untargeted hair lipidomic analysis and facilitate the use of hair in translational studies.

Assessment of silicone wristbands for monitoring personal exposure to chlorinated paraffins (C8-36): A pilot study.

Chlorinated paraffins (CPs) are a major environmental concern due to their ubiquitous presence in the environment. Since human exposure to CPs can significantly differ among individuals, it is essential to have an effective tool for monitoring personal exposure to CPs. In this pilot study, silicone wristbands (SWBs) were employed as a personal passive sampler to measure time-weighted average exposure to CPs. Twelve participants were asked to wear a pre-cleaned wristband for a week during the summer of 2022, and three field samplers (FSs) in different micro-environments were also deployed. The samples were then analyzed for CP homologs by LC-Q-TOFMS. In worn SWBs, the median concentrations of quantifiable CP classes were 19 ng/g wb, 110 ng/g wb, and 13 ng/g wb for ∑SCCPs, ∑MCCPs, and ∑LCCPs (C18-20), respectively. For the first time, lipid content is reported in worn SWBs, which could be a potential impact factor in the kinetics of the accumulation process for CPs. Results showed that micro-environments were key contributors to dermal exposure to CPs, while a few outliers suggested other sources of exposure. CP exposure via dermal contact showed an increased contribution and thus poses a nonnegligible potential risk to humans in daily life. Results presented here provide proof of concept of the use of SWBs as a cheap and non-invasive personal sampler in exposure studies.

A retrospective investigation of feather corticosterone in a highly contaminated white-tailed eagle (Haliaeetus albicilla) population.

Exposure to persistent organic pollutants (POPs), such as organochlorines (OCs) and polybrominated diphenyl ethers (PBDEs), is associated with adverse health effects in wildlife. Many POPs have been banned and consequently their environmental concentrations have declined. To assess both temporal trends of POPs and their detrimental impacts, raptors are extensively used as biomonitors due to their high food web position and high contaminant levels. White-tailed eagles (WTEs; Haliaeetus albicilla) in the Baltic ecosystem represent a sentinel species of environmental pollution, as they have suffered population declines due to reproductive failure caused by severe exposure to dichlorodiphenyltrichloroethane (DDT) and polychlorinated biphenyls (PCB) during the 1960s through 1980s. However, there is a lack of long-term studies that cover a wide range of environmental contaminants and their effects at the individual level. In this study, we used 135 pooled samples of shed body feathers collected in 1968-2012 from breeding WTE pairs in Sweden. Feathers constitute a temporal archive for substances incorporated into the feather during growth, including corticosterone, which is the primary avian glucocorticoid and a stress-associated hormone. Here, we analysed the WTE feather pools to investigate annual variations in feather corticosterone (fCORT), POPs (OCs and PBDEs), and stable carbon and nitrogen isotopes (SIs; dietary proxies). We examined whether the expected fluctuations in POPs affected fCORT (8-94 pg. mm-1) in the WTE pairs. Despite clear temporal declining trends in POP concentrations (p < 0.01), we found no significant associations between fCORT and POPs or SIs (p > 0.05 in all cases). Our results do not support fCORT as a relevant biomarker of contaminant-mediated effects in WTEs despite studying a highly contaminated population. However, although not detecting a relationship between fCORT, POP contamination and diet, fCORT represents a non-destructive and retrospective assessment of long-term stress physiology in wild raptors otherwise not readily available.

Hair as an alternative matrix to assess exposure of premature neonates to phthalate and alternative plasticizers in the neonatal intensive care unit.

Due to adverse health effects, di-(2-ethylhexyl) phthalate (DEHP), a plasticizer used to soften plastic medical devices (PMDs), was restricted, and gradually replaced by alternative plasticizers (APs). Up to this date, urine was the sole matrix studied for plasticizer exposure in neonates hospitalized in the neonatal intensive care unit (NICU), a population highly vulnerable to toxic effects of plasticizers. The primary aim of this study was to assess simultaneous measurement of phthalate and AP metabolites in neonatal scalp hair. In addition, we aimed to use this matrix to investigate exposure of premature neonates to plasticizers during their stay in the NICU. Hair samples in this study were collected from premature neonates and their mothers included in a prospective birth cohort study in a tertiary NICU at the Antwerp University Hospital (UZA), Belgium. Samples from premature neonates (n = 45) and their mothers (n = 107) as well as from control neonates (n = 24) and mothers (n = 29) were analyzed using liquid-chromatography coupled to tandem mass spectrometry. This is the first study reporting metabolites of phthalate and alternative plasticizers in neonatal hair samples as biomarkers for exposure to these plasticizers. Results showed that hair sampled from premature neonates after a NICU stay contained significantly higher metabolite concentrations of both phthalates (DEHP, DiBP, and DnBP; 9.0-2500, 9.3-2200, and 24.7-5300 ng/g), and alternative plasticizers (DEHA, DEHT, and TOTM; 38.8-3400, 127.5-5700, and 10.8-8700 ng/g) - when compared to healthy control neonates. Besides, DEHP and DEHT metabolite concentrations were significantly higher than in hair sampled from adult populations. In addition, prolonged NICU exposure to non-invasive respiratory support devices and gastric tubes was correlated with increased concentrations in hair samples, indicating accumulation of plasticizers in this alternative matrix. In conclusion, our data indicate that preterm neonates are still highly exposed to phthalate and alternative plasticizers during NICU stay, despite the EU Medical Devices Regulation.

Non-targeted proteomics reveals altered immune response in geographically distinct populations of green sea turtles (Chelonia mydas).

All seven species of sea turtle are facing increasing pressures from human activities that are impacting their health. Changes in circulating blood proteins of an individual, or all members of a population, can provide an early indicator of adverse health outcomes. Non-targeted measurement of all detectable proteins in a blood sample can indicate physiological changes. In the context of wildlife toxicology, this technique can provide a powerful tool for discovering biomarkers of chemical exposure and effect. This study presents a non-targeted examination of the protein abundance in sea turtle plasma obtained from three geographically distinct foraging populations of green turtles (Chelonia mydas) on the Queensland coast. Relative changes in protein expression between sites were compared, and potential markers of contaminant exposure were investigated. Blood plasma protein profiles were distinct between populations, with 85 out of the 116 identified proteins differentially expressed (p < 0.001). The most strongly dysregulated proteins were predominantly acute phase proteins, suggestive of differing immune status between the populations. The highest upregulation of known markers of immunotoxicity, such as pentraxin fusion and complement factor h, was observed in the Moreton Bay turtles. Forty-five different organohalogens were also measured in green turtle plasma samples as exposure to some organohalogens (e.g., polychlorinated biphenyls) has previously been identified as a cause for immune dysregulation in marine animals. The few detected organohalogens were at very low (pg/mL) concentrations in turtles from all sites, and are unlikely to be the cause of the proteome differences observed. However, the changes in protein expression may be indicative of exposure to other chemicals or environmental stressors. The results of this study provide important information about differences in protein expression between different populations of turtles, and guide future toxicological and health studies on east-Australian green sea turtles.

Metabolic signature of HepaRG cells exposed to ethanol and tumor necrosis factor alpha to study alcoholic steatohepatitis by LC-MS-based untargeted metabolomics.

Despite the high prevalence of alcoholic liver disease, its identification and characterization remain poor, especially in early stages such as alcoholic fatty liver disease and alcoholic steatohepatitis. This latter implies diagnostic difficulties, few therapeutic options and unclear mechanisms of action. To elucidate the metabolic alterations and pinpoint affected biochemical pathways, alcoholic steatohepatitis was simulated in vitro by exposing HepaRG cells to ethanol (IC10, 368 mM) and tumor necrosis factor alpha (TNF-α, 50 ng/mL) for 24 h. This combined exposure was compared to solely ethanol-exposed as well as -nonexposed cells. Four different metabolomics platforms were used combining liquid chromatography, high-resolution mass spectrometry and drift tube ion mobility to elucidate both intracellular and extracellular metabolic alterations. Some of the key findings include the influence of TNF-α in the upregulation of hepatic triglycerides and the downregulation of hepatic phosphatidylethanolamines and phosphatidylcholines. S-Adenosylmethionine showed to play a central role in the progression of alcoholic steatohepatitis. In addition, fatty acyl esters of hydroxy fatty acid (FAHFA)-containing triglycerides were detected for the first time in human hepatocytes and their alterations showed a potentially important role during the progression of alcoholic steatohepatitis. Ethoxylated phosphorylcholine was identified as a potential new biomarker of ethanol exposure.

Degradation products of resin-based materials detected in saliva in vivo.

OBJECTIVES: Dental composites remain under scrutiny regarding their (long-term) safety. In spite of numerous studies on the release of monomers both in vitro and in vivo, only limited quantitative data exist on the in vivo leaching of degradation products from monomers and additives. The aim of this observational study was for the first time to quantitatively and qualitatively monitor the release of parent compounds and their degradation products in saliva from patients undergoing multiple restorations. MATERIALS AND METHODS: Five patients in need of multiple large composite restorations (minimally 5 up to 28 restorations) due to wear (attrition, abrasion, and erosion) were included in the study, and they received adhesive restorative treatment according to the standard procedures in the university clinic for Restorative Dentistry. Saliva was collected at different time points, starting before the restoration up until 24 h after the treatment with composite restorations. Saliva extracts were analyzed by liquid chromatography-mass spectrometry. RESULTS: Leaching of monomers and degradation products was highest within 30 min after the placement of the restorations. The highest median concentrations of monomers were recorded for UDMA, BisEMA-3, and TEGDMA; yet, besides BisEMA-3 and TEGDMA, no monomers could be detected after 24 h. Mono- and demethacrylated degradation products remained present up to 24 h and concentrations were generally higher than those of monomers. In patients with multiple restorations, degradation products were still present in the sample taken before the next operation, several weeks after the previous operation. CONCLUSIONS: Exposure to residual monomers and degradation products occurs in the first hours after restoration. Monomers are present in saliva shortly after restoration, but degradation products can be detected weeks after the restoration confirming a long-term release. CLINICAL SIGNIFICANCE: Future research should focus more on the release of degradation products from monomers and additives from resin-based materials given their prolonged presence in saliva after restoration.

Metabolic Signature of Ethanol-Induced Hepatotoxicity in HepaRG Cells by Liquid Chromatography-Mass Spectrometry-Based Untargeted Metabolomics.

Alcoholic liver disease is highly prevalent but poorly identified and characterized, leading to knowledge gaps, which impairs early diagnosis. Excessive alcohol consumption is known to alter lipid metabolism, followed by progressive intracellular lipid accumulation, resulting in alcoholic fatty liver disease. In this study, HepaRG cells were exposed to ethanol at IC10 and 1/10 IC10 for 24 and 48 h. Metabolic alterations were investigated intra- and extracellularly with liquid chromatography-high-resolution mass spectrometry. Ion mobility was added as an extra separation dimension for untargeted lipidomics to improve annotation confidence. Distinctive patterns between exposed and control cells were consistently observed, with intracellular upregulation of di- and triglycerides, downregulation of phosphatidylcholines and phosphatidylethanolamines, sphingomyelins, and S-adenosylmethionine, among others. Several intracellular metabolic patterns could be related to changes in the extracellular environment, such as increased intracellular hydrolysis of sphingomyelins, leading to increased phosphorylcholine secretion. Carnitines showed alterations depending on the size of their carbon chain, which highlights the interplay between ß-oxidation in mitochondria and peroxisomes. Potential new biomarkers of ethanol-induced hepatotoxicity have been observed, such as ceramides with a sphingadienine backbone, octanoylcarnitine, creatine, acetylcholine, and ethoxylated phosphorylcholine. The combination of the metabolic fingerprint and footprint enabled a comprehensive investigation of the pathophysiology behind ethanol-induced hepatotoxicity.

Guidelines and considerations for building multidimensional libraries for untargeted MS-based metabolomics.

INTRODUCTION: Feature annotation is crucial in untargeted metabolomics but remains a major challenge. The large pool of metabolites collected under various instrumental conditions is underrepresented in publicly available databases. Retention time (RT) and collision cross section (CCS) measurements from liquid chromatography ion mobility high-resolution mass spectrometers can be employed in addition to MS/MS spectra to improve the confidence of metabolite annotation. Recent advancements in machine learning focus on improving the accuracy of predictions for CCS and RT values. Therefore, high-quality experimental data are crucial to be used either as training datasets or as a reference for high-confidence matching. METHODS: This manuscript provides an easy-to-use workflow for the creation of an in-house metabolite library, offers an overview of alternative solutions, and discusses the challenges and advantages of using open-source software. A total of 100 metabolite standards from various classes were analyzed and subjected to the described workflow for library generation. RESULTS AND DISCUSSION: The outcome was an open-access available NIST format metabolite library (.msp) with multidimensional information. The library was used to evaluate CCS prediction tools, MS/MS spectra heterogeneities (e.g., multiple adducts, in-source fragmentation, radical fragment ions using collision-induced dissociation), and the reporting of RT.

Neurodevelopmental toxicity assessment of flame retardants using a human DNT in vitro testing battery.

Due to their neurodevelopmental toxicity, flame retardants (FRs) like polybrominated diphenyl ethers are banned from the market and replaced by alternative FRs, like organophosphorus FRs, that have mostly unknown toxicological profiles. To study their neurodevelopmental toxicity, we evaluated the hazard of several FRs including phased-out polybrominated FRs and organophosphorus FRs: 2,2',4,4'-tetrabromodiphenylether (BDE-47), 2,2',4,4',5-pentabromodiphenylether (BDE-99), tetrabromobisphenol A, triphenyl phosphate, tris(2-butoxyethyl) phosphate and its metabolite bis-(2-butoxyethyl) phosphate, isodecyl diphenyl phosphate, triphenyl isopropylated phosphate, tricresyl phosphate, tris(1,3-dichloro-2-propyl) phosphate, tert-butylphenyl diphenyl phosphate, 2-ethylhexyl diphenyl phosphate, tris(1-chloroisopropyl) phosphate, and tris(2-chloroethyl) phosphate. Therefore, we used a human cell-based developmental neurotoxicity (DNT) in vitro battery covering a large variety of neurodevelopmental endpoints. Potency according to the respective most sensitive benchmark concentration (BMC) across the battery ranked from <1 µM (5 FRs), 1<10 µM (7 FRs) to the >10 µM range (3 FRs). Evaluation of the data with the ToxPi tool revealed a distinct ranking (a) than with the BMC and (b) compared to the ToxCast data, suggesting that DNT hazard of these FRs is not well predicted by ToxCast assays. Extrapolating the DNT in vitro battery BMCs to human FR exposure via breast milk suggests low risk for individual compounds. However, it raises a potential concern for real-life mixture exposure, especially when different compounds converge through diverse modes-of-action on common endpoints, like oligodendrocyte differentiation in this study. This case study using FRs suggests that human cell-based DNT in vitro battery is a promising approach for neurodevelopmental hazard assessment and compound prioritization in risk assessment.