The genes mgtE and spoVG are involved in zinc tolerance of Staphylococcus aureus.

Metals are essential for all living organisms, but the type of metal and its concentration determines its action. Even low concentrations of metals may have toxic effects on organisms and therefore exhibit antimicrobial activities. In this study, we investigate the evolutionary adaptation processes of Staphylococcus aureus to metals and common genes for metal tolerance. Laboratory and clinical isolates were treated with manganese, cobalt, zinc, or nickel metal salts to generate growth-adapted mutants. After growth in medium supplemented with zinc, whole-genome sequencing identified, among others, two genes, mgtE (SAUSA300_0910), a putative magnesium transporter and spoVG (SAUSA300_0475), a global transcriptional regulator, as hot spots for stress-induced single-nucleotide polymorphisms (SNPs). SNPs in mgtE were also detected in mutants treated with high levels of cobalt or nickel salts. To investigate the effect of these genes on metal tolerance, deletion mutants and complementation strains in an S. aureus USA300 LAC* laboratory strain were generated. Both, the mgtE and spoVG deletion strains were more tolerant to cobalt, manganese, and zinc. The mgtE mutant was also more tolerant to nickel exposure. Inductively coupled plasma mass spectrometry analysis demonstrated that the mgtE deletion mutant accumulated less intracellular zinc than the wild type, explaining increased tolerance. From these results, we conclude that mgtE gene inactivation increases zinc tolerance presumably due to reduced uptake of zinc. For the SpoVG mutant, no direct effect on the intracellular zinc concentration was detected, indicating toward different pathways to increase tolerance. Importantly, inactivation of these genes offers a growth advantage in environments containing certain metals, pointing toward a common tolerance mechanism. IMPORTANCE: Staphylococcus aureus is an opportunistic pathogen causing tremendous public health burden and high mortality in invasive infections. Treatment is becoming increasingly difficult due to antimicrobial resistances. The use of metals in animal husbandry and aquaculture to reduce bacterial growth and subsequent acquisition of metal resistances has been shown to co-select for antimicrobial resistance. Therefore, understanding adaptive mechanisms that help S. aureus to survive metal exposure is essential. Using a screening approach, we were able to identify two genes encoding the transporter MgtE and the transcriptional regulator SpoVG, which conferred increased tolerance to specific metals such as zinc when inactivated. Further testing showed that the deletion of mgtE leads to reduced intracellular zinc levels, suggesting a role in zinc uptake. The accumulation of mutations in these genes when exposed to other metals suggests that inactivation of these genes could be a common mechanism for intrinsic tolerance to certain metals.

Influence of Silver Fiber Morphology on the Dose-Response Relationship and Enrichment in Daphnia magna Studied by Elemental Imaging with LA-ICP-TOF-MS.

This study aims to enhance the understanding of the environmental risks associated with nanomaterials, particularly nanofibers. Previous research suggested that silver fibers exhibit higher toxicity (EC50/48h 1.6-8.5 µg/L) compared to spherical silver particles (EC50/48h 43 µg/L). To investigate the hypothesis that toxicity is influenced by the morphology and size of nanomaterials, various silver nanofibers with different dimensions (length and diameter) were selected. The study assessed their toxicity toward Daphnia magna using the 48 h immobilization assay. The EC50 values for the different fibers ranged from 122 to 614 µg/L. Subsequently, the study quantified the uptake and distribution of two representative nanofibers in D. magna neonates by employing digestion and imaging mass spectrometry in the form of laser-ablation-ICP-MS. A novel sample preparation method was utilized, allowing the analysis of whole, intact daphnids, which facilitated the localization of silver material and prevented artifacts. The results revealed that, despite the similar ecotoxicity of the silver fibers, the amount of silver associated with the neonates differed by a factor of 2-3. However, both types of nanofibers were primarily found in the gut of the organisms. In conclusion, the findings of this study do not support the expectation that the morphology or size of silver materials affect their toxicity to D. magna.

Post column infusion of an internal standard into LC-FT-ICR MS enables semi-quantitative comparison of dissolved organic matter in original samples.

Ultrahigh resolution mass spectrometry hyphenated with liquid chromatography (LC) is an emerging tool to explore the isomeric composition of dissolved organic matter (DOM). However, matrix effects limit the potential for semi-quantitative comparison of DOM molecule abundances across samples. We introduce a post-column infused internal standard (PCI-IS) for reversed-phase LC-FT-ICR MS measurements of DOM and systematically evaluate matrix effects, detector linearity and the precision of mass peak intensities. Matrix effects for model compounds spiked into freshwater DOM samples ranging from a headwater stream to a major river were reduced by 5-10% for PCI-IS corrected mass peak intensities as compared to raw (i.e., untransformed) intensities. A linear regression of PCI-IS corrected DOM mass peak intensities across a typical DOM concentration range (2-15 mg dissolved organic carbon L-1) in original, non-extracted freshwater samples demonstrates excellent linearity of the detector response (r2 > 0.9 for 98% of detected molecular formulas across retention times). Importantly, PCI-IS could compensate for 80% of matrix effects across an environmental gradient of DOM composition from groundwater to surface water. This enabled studying the ionization efficiency of DOM isomers and linking the observed differences to the biogeochemical sources. With PCI-IS original, non-extracted DOM samples can be analysed by LC-FT-ICR MS without carbon load adjustment, and mass peak intensities can be reliably used to semi-quantitatively compare isomer abundances between compositionally similar DOM samples.

Hotspots of Floating Plastic Particles across the North Pacific Ocean.

The pollution of the marine environment with plastic debris is expected to increase, where ocean currents and winds cause their accumulation in convergence zones like the North Pacific Subtropical Gyre (NPSG). Surface-floating plastic (>330 µm) was collected in the North Pacific Ocean between Vancouver (Canada) and Singapore using a neuston catamaran and identified by Fourier-transform infrared spectroscopy (FT-IR). Baseline concentrations of 41,600-102,700 items km-2 were found, dominated by polyethylene and polypropylene. Higher concentrations (factors 4-10) of plastic items occurred not only in the NPSG (452,800 items km-2) but also in a second area, the Papaha̅naumokua̅kea Marine National Monument (PMNM, 285,200 items km-2). This second maximum was neither reported previously nor predicted by the applied ocean current model. Visual observations of floating debris (>5 cm; 8-2565 items km-2 and 34-4941 items km-2 including smaller "white bits") yielded similar patterns of baseline pollution (34-3265 items km-2) and elevated concentrations of plastic debris in the NPSG (67-4941 items km-2) and the PMNM (295-3748 items km-2). These findings suggest that ocean currents are not the only factor provoking plastic debris accumulation in the ocean. Visual observations may be useful to increase our knowledge of large-scale (micro)plastic pollution in the global oceans.

Should Transformation Products Change the Way We Manage Chemicals?

When chemical pollutants enter the environment, they can undergo diverse transformation processes, forming a wide range of transformation products (TPs), some of them benign and others more harmful than their precursors. To date, the majority of TPs remain largely unrecognized and unregulated, particularly as TPs are generally not part of routine chemical risk or hazard assessment. Since many TPs formed from oxidative processes are more polar than their precursors, they may be especially relevant in the context of persistent, mobile, and toxic (PMT) and very persistent and very mobile (vPvM) substances, which are two new hazard classes that have recently been established on a European level. We highlight herein that as a result, TPs deserve more attention in research, chemicals regulation, and chemicals management. This perspective summarizes the main challenges preventing a better integration of TPs in these areas: (1) the lack of reliable high-throughput TP identification methods, (2) uncertainties in TP prediction, (3) inadequately considered TP formation during (advanced) water treatment, and (4) insufficient integration and harmonization of TPs in most regulatory frameworks. A way forward to tackle these challenges and integrate TPs into chemical management is proposed.

Revisiting Disinfection Byproducts with Supercritical Fluid Chromatography-High Resolution-Mass Spectrometry: Identification of Novel Halogenated Sulfonic Acids in Disinfected Drinking Water.

High resolution mass spectrometry (HRMS) coupled to either gas chromatography or reversed-phase liquid chromatography is the generic method to identify unknown disinfection byproducts (DBPs) but can easily overlook their highly polar fractions. In this study, we applied an alternative chromatographic separation method, supercritical fluid chromatography-HRMS, to characterize DBPs in disinfected water. In total, 15 DBPs were tentatively identified for the first time as haloacetonitrilesulfonic acids, haloacetamidesulfonic acids, and haloacetaldehydesulfonic acids. Cysteine, glutathione, and p-phenolsulfonic acid were found as precursors during lab-scale chlorination, with cysteine providing the highest yield. A mixture of the labeled analogues of these DBPs was prepared by chlorination of 13C3-15N-cysteine and analyzed using nuclear magnetic resonance spectroscopy for structural confirmation and quantification. A total of 6 drinking water treatment plants utilizing various source waters and treatment trains produced sulfonated DBPs upon disinfection. Those were widespread in the tap water of 8 cities across Europe, with estimated concentrations up to 50 and 800 ng/L for total haloacetonitrilesulfonic acids and haloacetaldehydesulfonic acids, respectively. Up to 850 ng/L haloacetonitrilesulfonic acids were found in 3 public swimming pools. Considering the stronger toxicity of haloacetonitriles, haloacetamides, and haloacetaldehydes than the regulated DBPs, these newly found sulfonic acid derivatives may also pose a health risk.

Uptake and Biotransformation of the Tire Rubber-derived Contaminants 6-PPD and 6-PPD Quinone in the Zebrafish Embryo (Danio rerio).

N-(1,3-Dimethylbutyl)-N'-phenyl-p-phenylenediamine (6-PPD) is a widely used antioxidant in tire rubber known to enter the aquatic environment via road runoff. The associated transformation product (TP) 6-PPD quinone (6-PPDQ) causes extreme acute toxicity in some fish species (e.g., coho salmon). To interpret the species-specific toxicity, information about biotransformation products of 6-PPDQ would be relevant. This study investigated toxicokinetics of 6-PPD and 6-PPDQ in the zebrafish embryo (ZFE) model. Over 96 h of exposure, 6-PPD and 6-PPDQ accumulated in the ZFE with concentration factors ranging from 140 to 2500 for 6-PPD and 70 to 220 for 6-PPDQ. A total of 22 TPs of 6-PPD and 12 TPs of 6-PPDQ were tentatively identified using liquid chromatography coupled to high-resolution mass spectrometry. After 96 h of exposure to 6-PPD, the TPs of 6-PPD comprised 47% of the total peak area (TPA), with 4-hydroxydiphenylamine being the most prominent in the ZFE. Upon 6-PPDQ exposure, >95% of 6-PPDQ taken up in the ZFE was biotransformed, with 6-PPDQ + O + glucuronide dominating (>80% of the TPA). Among other TPs of 6-PPD, a reactive N-phenyl-p-benzoquinone imine was found. The knowledge of TPs of 6-PPD and 6-PPDQ from this study may support biotransformation studies in other organisms.

The effect of root hairs on exudate composition: a comparative non-targeted metabolomics approach.

Root exudation is a major pathway of organic carbon input into soils. It affects soil physical properties, element solubility as well as speciation, and impacts the microbial community in the rhizosphere. Root exudates contain a large number of primary and secondary plant metabolites, and the amount and composition are highly variable depending on plant species and developmental stage. Detailed information about exudate composition will allow for a better understanding of exudate-driven rhizosphere processes and their feedback loops. Although non-targeted metabolomics by high-resolution mass spectrometry is an established tool to characterize root exudate composition, the extent and depth of the information obtained depends strongly on the analytical approach applied. Here, two genotypes of Zea mays L., differing in root hair development, were used to compare six mass spectrometric approaches for the analysis of root exudates. Reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography combined with time-of-flight mass spectrometry (LC-TOF-MS), as well as direct infusion Fourier-transform ion cyclotron resonance mass spectrometry (DI-FT-ICR-MS), were applied with positive and negative ionization mode. By using the same statistical workflow, the six approaches resulted in different numbers of detected molecular features, ranging from 176 to 889, with a fraction of 48 to 69% of significant features (fold change between the two genotypes of > 2 and p-value < 0.05). All approaches revealed the same trend between genotypes, namely up-regulation of most metabolites in the root hair defective mutant (rth3). These results were in agreement with the higher total carbon and nitrogen exudation rate of the rth3-mutant as compared to the corresponding wild-type maize (WT). However, only a small fraction of features were commonly found across the different analytical approaches (20-79 features, 13-31% of the rth3-mutant up-regulated molecular formulas), highlighting the need for different mass spectrometric approaches to obtain a more comprehensive view into the composition of root exudates. In summary, 111 rth3-mutant up-regulated compounds (92 different molecular formulas) were detected with at least two different analytical approaches, while no WT up-regulated compound was found by both, LC-TOF-MS and DI-FT-ICR-MS. Zea mays L. exudate features obtained with multiple analytical approaches in our study were matched against the metabolome database of Zea mays L. (KEGG) and revealed 49 putative metabolites based on their molecular formula.

Zebrafish Oatp1d1 Acts as a Cellular Efflux Transporter of the Anionic Herbicide Bromoxynil.

Toxicokinetics (TK) of ionic compounds in the toxico-/pharmacological model zebrafish embryo (Danio rerio) depend on absorption, distribution, metabolism, and elimination (ADME) processes. Previous research indicated involvement of transport proteins in the TK of the anionic pesticide bromoxynil in zebrafish embryos. We here explored the interaction of bromoxynil with the organic anion-transporting polypeptide zebrafish Oatp1d1. Mass spectrometry imaging revealed accumulation of bromoxynil in the gastrointestinal tract of zebrafish embryos, a tissue known to express Oatp1d1. In contrast to the Oatp1d1 reference substrate bromosulfophthalein (BSP), which is actively taken up by transfected HEK293 cells overexpressing zebrafish Oatp1d1, those cells accumulated less bromoxynil than empty vector-transfected control cells. This indicates cellular efflux of bromoxynil by Oatp1d1. This was also seen for diclofenac but not for carbamazepine, examined for comparison. Correspondingly, internal concentrations of bromoxynil and diclofenac in the zebrafish embryo were increased when coexposed with BSP, inhibiting the activities of various transporter proteins, including Oatp1d1. The effect of BSP on accumulation of bromoxynil and diclofenac was enhanced in further advanced embryo stages, indicating increased efflux activity in those stages. An action of Oatp1d1 as an efflux transporter of ionic environmental compounds in zebrafish embryos should be considered in future TK assessments.

Discovery of Polar Ozonation Byproducts via Direct Injection of Effluent Organic Matter with Online LC-FT-ICR-MS.

Effluent organic matter (EfOM), a major ozone consumer during wastewater ozonation, is a complex mixture of natural and anthropogenic organic molecules. Ozonation of EfOM adds to molecular complexity by introducing polar and potentially mobile ozonation byproducts (OBPs). Currently, nontargeted direct infusion (DI) ultrahigh resolution mass spectrometry (e.g. FT-ICR-MS) is used to study OBPs but requires sample extraction, limiting the accessible polarity range of OBPs. To better understand the impact of ozonation on EfOM and the formation of polar OBPs, nonextracted effluent was analyzed by direct injection onto a reversed-phase liquid chromatography system (RP-LC) online hyphenated with an FT-ICR-MS. Over four times more OBPs were detected in nonextracted EfOM compared to effluent extracted with solid phase extraction and measured with DI-FT-ICR-MS (13817 vs 3075). Over 1500 highly oxygenated OBPs were detected exclusively in early eluting fractions of nonextracted EfOM, indicating polar OBPs. Oxygenation of these newly discovered OBPs is higher than previously found, with an average molecular DBE-O value of -3.3 and O/C ratio of 0.84 in the earliest eluting OBP fractions. These polar OBPs are consistently lost during extraction but may play an important role in understanding the environmental impact of ozonated EfOM. Moreover, 316 molecular formulas classified as nonreactive to ozone in DI-FT-ICR-MS can be identified with LC-FT-ICR-MS as isomers with varying degrees of reactivity, providing for the first time experimental evidence of differential reactivity of complex organic matter isomers with ozone.

Sorption and Mobility of Charged Organic Compounds: How to Confront and Overcome Limitations in Their Assessment.

Permanently charged and ionizable organic compounds (IOC) are a large and diverse group of compounds belonging to many contaminant classes, including pharmaceuticals, pesticides, industrial chemicals, and natural toxins. Sorption and mobility of IOCs are distinctively different from those of neutral compounds. Due to electrostatic interactions with natural sorbents, existing concepts for describing neutral organic contaminant sorption, and by extension mobility, are inadequate for IOC. Predictive models developed for neutral compounds are based on octanol-water partitioning of compounds (Kow) and organic-carbon content of soil/sediment, which is used to normalize sorption measurements (KOC). We revisit those concepts and their translation to IOC (Dow and DOC) and discuss compound and soil properties determining sorption of IOC under water saturated conditions. Highlighting possible complementary and/or alternative approaches to better assess IOC mobility, we discuss implications on their regulation and risk assessment. The development of better models for IOC mobility needs consistent and reliable sorption measurements at well-defined chemical conditions in natural porewater, better IOC-, as well as sorbent characterization. Such models should be complemented by monitoring data from the natural environment. The state of knowledge presented here may guide urgently needed future investigations in this field for researchers, engineers, and regulators.

Thermophilic archaea activate butane via alkyl-coenzyme M formation.

The anaerobic formation and oxidation of methane involve unique enzymatic mechanisms and cofactors, all of which are believed to be specific for C1-compounds. Here we show that an anaerobic thermophilic enrichment culture composed of dense consortia of archaea and bacteria apparently uses partly similar pathways to oxidize the C4 hydrocarbon butane. The archaea, proposed genus 'Candidatus Syntrophoarchaeum', show the characteristic autofluorescence of methanogens, and contain highly expressed genes encoding enzymes similar to methyl-coenzyme M reductase. We detect butyl-coenzyme M, indicating archaeal butane activation analogous to the first step in anaerobic methane oxidation. In addition, Ca. Syntrophoarchaeum expresses the genes encoding ß-oxidation enzymes, carbon monoxide dehydrogenase and reversible C1 methanogenesis enzymes. This allows for the complete oxidation of butane. Reducing equivalents are seemingly channelled to HotSeep-1, a thermophilic sulfate-reducing partner bacterium known from the anaerobic oxidation of methane. Genes encoding 16S rRNA and methyl-coenzyme M reductase similar to those identifying Ca. Syntrophoarchaeum were repeatedly retrieved from marine subsurface sediments, suggesting that the presented activation mechanism is naturally widespread in the anaerobic oxidation of short-chain hydrocarbons.

Online Counter Gradient LC-FT-ICR-MS Enables Detection of Highly Polar Natural Organic Matter Fractions.

Natural organic matter (NOM) is a highly complex mixture of natural organic molecules. The recent developments in NOM molecular characterization methods have shown that ESI-FT-ICR hyphenated with liquid chromatography (LC) is a promising approach to also obtain chemical information (such as polarity and molecular size) about NOM molecules. However, due to changing solvent composition during gradient elution in LC-FT-ICR-MS, ionization conditions also change throughout the chromatographic separation process. In this study, we applied a post-LC column counter gradient (CG) to ensure stable solvent conditions for transient ESI-MS signals. Suwanee River Fulvic Acid (SRFA) standard and a peat pore water were used as representative dissolved NOM samples for method development and validation. Our results show that in polar NOM fractions (which elute with <50% methanol) the TIC intensity and number of assigned molecular formulas were increased by 48% and 20%, as compared to the standard gradient (SG) method. Further application of a Q-isolation and selective ion accumulation for low abundance fractions revealed over 3 times more molecular formulas (especially for CHNO, CHOS, CHNOS formula classes) than in full scan mode. The number of detected highly polar NOM compounds (with elemental ratios H/C < 1, O/C > 0.6) were more than 20 times larger for CG-LC mode as compared to direct infusion (DI) (5715 vs 266 MF). We conclude that the application of a postcolumn counter gradient in LC-FT-ICR-MS analyses of NOM offers novel insight into the most polar fractions of NOM which are inaccessible in conventional DI measurements.

Prenatal paraben exposure and atopic dermatitis-related outcomes among children.

BACKGROUND: Parabens, widely used as preservatives in cosmetics, foods, and other consumer products, are suspected of contributing to allergy susceptibility. The detection of parabens in the placenta or amniotic fluid raised concerns about potential health consequences for the child. Recently, an increased asthma risk following prenatal exposure has been reported. Here, we investigated whether prenatal paraben exposure can influence the risk for atopic dermatitis (AD). METHODS: 261 mother-child pairs of the German mother-child study LINA were included in this analysis. Eight paraben species were quantified in maternal urine obtained at gestational week 34. According to the parental report of physician-diagnosed AD from age 1 to 8 years, disease onset, and persistence, childhood AD was classified into four different phenotypes. RESULTS: 4.6% (n = 12) and 12.3% (n = 32) of the children were classified as having very early-onset AD (until age two) either with or without remission, 11.9% (n = 31) as early-onset (after age two), and 3.1% (n = 8) as childhood-onset AD (after age six). Exposure to ethylparaben and n-butylparaben was associated with an increased risk to develop very early-onset AD without remission (EtP: adj.OR/95% CI:1.44/1.04-2.00,nBuP:adj.OR/95% CI:1.95/1.22-3.12). The effects of both parabens were predominant in children without a history of maternal AD and independent of children's sex. CONCLUSION: Prenatal EtP or nBuP exposure may increase children's susceptibility for persistent AD with disease onset at very early age. This association was particularly pronounced in children without a history of maternal AD, indicating that children without a genetic predisposition are more susceptible to paraben exposure.

Ruthenium red: a highly efficient and versatile cell staining agent for single-cell analysis using inductively coupled plasma time-of-flight mass spectrometry.

Staining of biological cells with heavy metals can increase their visibility in mass spectrometry. In this study, the potential of ruthenium red (RR) as a staining agent for single-cell analysis by inductively coupled plasma time-of-flight mass spectrometry (SC-ICP-TOF-MS) is explored using two different yeast strains and one algal species. Time-of-flight mass spectrometry allows the simultaneous detection of Ru and multiple intrinsic elements in single cells. Ru has a better correlation with Mg than with P in Saccharomyces cerevisiae (S. cerevisiae) cells. For the three tested strains, the staining efficiency of RR exceeded 96%; the staining strengths were 30-32 ag µm-2 for the yeast cells and 59 ag µm-2 for the algal cells. By deriving the cell volume of single cells from their Ru mass, the concentration of Mg and P in individual cells of S. cerevisiae can be calculated. Elemental concentrations of Mg and P were highly variable in the cell individuals, with their 25-75 percentile values of 0.10-0.19 and 0.76-2.07 fg µm-3, respectively. RR staining has several advantages: it is fast, does not affect cell viability and is highly efficient. Provided that the shape of the individual cells of a culture is similar, Ru staining allows the elemental content to be directly correlated with the cell volume to accurately calculate the intracellular concentration of target elements in single cells. Therefore, RR can be a promising cell staining agent for future application in SC-ICP-TOF-MS research.

Organic Markers of Tire and Road Wear Particles in Sediments and Soils: Transformation Products of Major Antiozonants as Promising Candidates.

Tire and road wear particles (TRWPs) are one of the main sources of particulate traffic emissions, but measured data on TRWP contents in the environment are scarce. This study aims at identifying organic compounds suitable as quantitative markers for TRWPs by a tiered multistep selection process involving nontarget screening and subsequent identification by liquid-chromatography high-resolution mass spectrometry. Starting from several thousands of signals recorded in the extract of tire particles, the rigorous selection process considered source specificity, tendency of leaching, analytical sensitivity and precision, and stability during aging. It led to three transformation products of N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6-PPD) as the most suitable marker candidates: N-formyl-6-PPD, hydroxylated N-1,3-dimethylbutyl-N-phenyl quinone diimine, and 6-PPD-quinone. A linear response in standard addition experiments with tire particles and the correlation with TRWP contents in a diverse set of environmental samples imply that these compounds are promising candidates as markers for the quantification of TRWPs. Organic markers for TRWP contents in the environment would allow TRWP quantification with the traditional tandem MS (LC-MS/MS) equipment of an organic trace analytical laboratory and, thus, allow easy generation of data on TRWP occurrence in sediments and soils and other environmental matrices.

Abiotic Transformation of Lamotrigine by Redox-Active Mineral and Phenolic Compounds.

The anticonvulsant drug lamotrigine is a recalcitrant environmental pollutant. It was detected in drinking water, surface water, reclaimed wastewater, arable soils, and even in edible crops. In this work, we studied the mechanisms of lamotrigine transformation by a common redox soil mineral, birnessite, in a single-solute system and in bisolute systems with vanillic acid or o-methoxyphenol. In the single-solute system, 28% of lamotrigine was transformed and 14 transformation products (TPs) were identified. Based on a detailed analysis of the TPs, we suggested that lamotrigine is transformed mainly by oxidation, addition, and dechlorination reactions. In the bisolute systems, the redox-active phenolic compounds enhanced the elimination and transformation of lamotrigine. Vanillic acid was more efficient, generating 92% transformation of lamotrigine (58 TPs were identified), whereas o-methoxyphenol induced 48% transformation (35 TPs were identified). In the bisolute system with phenolic compounds, lamotrigine has possibly been transformed mainly via addition reactions with phenolic compounds and their oxidation products (protocatechuic acid, quinone, and oligomers). Thus, masses of the formed TPs were elevated as compared to the parent compound. The current study demonstrates the important role of redox-active minerals and naturally occurring phenolic compounds in abiotic removal and transformation of a recalcitrant environmental pollutant.

Effective processing and evaluation of chemical imaging data with respect to morphological features of the zebrafish embryo.

A workflow was developed and implemented in a software tool for the automated combination of spatially resolved laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) data and data on the morphology of the biological tissue. Making use of a recently published biological annotation software, FishImager automatically assigns the biological feature as regions of interest (ROIs) and overlays them with the quantitative LA-ICP-MS data. Furthermore, statistical tools including cluster algorithms can be applied to the elemental intensity data and directly compared with the ROIs. This is effectively visualized in heatmaps. This allows gaining statistical significance on distribution and co-localization patterns. Finally, the biological functions of the assigned ROIs can then be easily linked with elemental distributions. We demonstrate the versatility of FishImager with quantitative LA-ICP-MS data of the zebrafish embryo tissue. The distribution of natural elements and xenobiotics is analyzed and discussed. With the help of FishImager, it was possible to identify compartments affected by toxicity effects or biological mechanisms to eliminate the xenobiotic. The presented workflow can be used for clinical and ecotoxicological testing, for example. Ultimately, it is a tool to simplify and reproduce interpretations of imaging LA-ICP-MS data in many applications.

Yolk Sac of Zebrafish Embryos as Backpack for Chemicals?

The zebrafish embryo (Danio rerio) has developed into one of the most important nonsentient animal models for the hazard assessments of chemicals, but the processes governing its toxicokinetics (TK) are poorly understood. This study compares the uptake of seven test compounds into the embryonic body and the yolk sac of the zebrafish embryo using TK experiments, a dialysis approach, thermodynamic calculations, and kinetic modeling. Experimental data show that between 95% (4-iodophenol) and 67% (carbamazepine) of the total internal amount in 26 h post fertilization (hpf) embryos and between 80 and 49% in 74 hpf embryos were found in the yolk. Thus, internal concentrations determined for the whole embryo overestimate the internal concentration in the embryonic body: for the compounds of this study, up to a factor of 5. Partition coefficients for the embryonic body and a one-compartment model with diffusive exchange were calculated for the neutral test compounds and agreed reasonably with the experimental data. For prevalently ionic test compounds at exposure pH (bromoxynil, paroxetine), however, the extent and the speed of uptake were low and could not be modeled adequately. A better understanding of the TK of ionizable test compounds is essential to allow assessment of the validity of this organismic test system for ionic test compounds.

Improved Understanding of Dissolved Organic Matter Processing in Freshwater Using Complementary Experimental and Machine Learning Approaches.

Dissolved organic matter plays an important role in aquatic ecosystems and poses a major problem for drinking water production. However, our understanding of DOM reactivity in natural systems is hampered by its complex molecular composition. Here, we used Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) and data from two independent studies to disentangle DOM reactivity based on photochemical and microbial-induced transformations. Robust correlations of FT-ICR-MS peak intensities with chlorophyll a and solar irradiation were used to define 9 reactivity classes for 1277 common molecular formulas. Germany's largest drinking water reservoir was sampled for 1 year, and DOM processing in stratified surface waters could be attributed to photochemical transformations during summer months. Microbial DOM alterations could be distinguished based on correlation coefficients with chlorophyll a and often shared molecular features (elemental ratios and mass) with photoreactive compounds. In particular, many photoproducts and some microbial products were identified as potential precursors of disinfection byproducts. Molecular DOM features were used to further predict molecular reactivity for the remaining compounds in the data set based on a random forest model. Our method offers an expandable classification approach to integrate the reactivity of DOM from specific environments and link it to molecular properties and chemistry.