Detection and Exclusion of False-Positive Molecular Formula Assignments via Mass Error Distributions in UHR Mass Spectra of Natural Organic Matter.

Ultrahigh resolution mass spectrometry (UHRMS) routinely detects and identifies thousands of mass peaks in complex mixtures, such as natural organic matter (NOM) and petroleum. The assignment of several chemically plausible molecular formulas (MFs) for a single accurate mass still poses a major problem for the reliable interpretation of NOM composition in a biogeochemical context. Applying sensible chemical rules for MF validation is often insufficient to eliminate multiple assignments (MultiAs)─especially for mass peaks with low abundance or if ample heteroatoms or isotopes are included - and requires manual inspection or expert judgment. Here, we present a new approach based on mass error distributions for the identification of true and false assignments among MultiAs. To this end, we used the mass error in millidalton (mDa), which was superior to the commonly used relative mass error in ppm. We developed an automatic workflow to group MultiAs based on their shared formula units and Kendrick mass defect values and to evaluate the mass error distribution. In this way, the number of valid assignments of chlorinated disinfection byproducts was increased by 8-fold as compared to only applying 37Cl/35Cl isotope ratio filters. Likewise, phosphorus-containing MFs can be differentiated against chlorine-containing MFs with high confidence. Further, false assignments of highly aromatic sulfur-containing MFs ("black sulfur") to sodium adducts in negative ionization mode can be excluded by applying our approach. Overall, MFs for mass peaks that are close to the detection limit or where naturally occurring isotopes are rare (e.g., 15N) or absent (e.g., P and F) can now be validated, substantially increasing the reliability of MF assignments and broadening the applicability of UHRMS analysis to even more complex samples and processes.

A Temporal Graph Model to Predict Chemical Transformations in Complex Dissolved Organic Matter.

Dissolved organic matter (DOM) is a complex mixture of thousands of natural molecules that undergo constant transformation in the environment, such as sunlight induced photochemical reactions. Despite molecular level resolution from ultrahigh resolution mass spectrometry (UHRMS), trends of mass peak intensities are currently the only way to follow photochemically induced molecular changes in DOM. Many real-world relationships and temporal processes can be intuitively modeled using graph data structures (networks). Graphs enhance the potential and value of AI applications by adding context and interconnections allowing the uncovering of hidden or unknown relationships in data sets. We use a temporal graph model and link prediction to identify transformations of DOM molecules in a photo-oxidation experiment. Our link prediction algorithm simultaneously considers educt removal and product formation for molecules linked by predefined transformation units (oxidation, decarboxylation, etc.). The transformations are further weighted by the extent of intensity change and clustered on the graph structure to identify groups of similar reactivity. The temporal graph is capable of identifying relevant molecules subject to similar reactions and enabling to study their time course. Our approach overcomes previous data evaluation limitations for mechanistic studies of DOM and leverages the potential of temporal graphs to study DOM reactivity by UHRMS.

Data evaluation strategy for identification of key molecular formulas in dissolved organic matter as proxies for biogeochemical reactivity based on abundance differences from ultrahigh resolution mass spectrometry.

The molecular composition of dissolved organic matter (DOM) is of relevance for global carbon cycling and important for drinking water processing also. The detection of variation of DOM composition as function of time and space from a methodological viewpoint is essential to observe DOM processing and was addressed so far. High resolution concerning DOM quality was achieved with Fourier-transform ion cyclotron resonance mass spectrometry (FTICR-MS). However almost none of the existing FTICR-MS data sets were evaluated addressing the fate of single mass features / molecular formulas (MFs) abundance during experiments. In contrast to former studies we analyze the function of MF abundance of time and space for such MFs which are present in all samples and which were formerly claimed as recalcitrant in not all but a great number of studies. For the first time the reactivity of MFs was directly compared by their abundance differences using a simple equation, the relative intensity difference (δRI). Search strategies to find out the maximum δRI values are introduced. The corresponding MFs will be regarded as key MFs (KEY-MFs). In order to test this new approach data from a recent photo degradation experiment were combined with monitoring surveys conducted in two drinking water reservoirs. The δRI values varied over one order of magnitude (more than five-fold). MFs like C9H12O6 and C10H14O6 revealed high biogeochemical reactivity as photo products. Some of the KEY-MFs were identical with MFs identified as disinfection byproducts precursors in recent studies. Other KEY-MFs were oxygen-rich and relatively unsaturated (poly-phenol-like) and hence relevant to flocculation procedures.

Lagrangian profiles of riverine autotrophy, organic matter transformation, and micropollutants at extreme drought.

On their way from inland to the ocean, flowing water bodies, their constituents and their biotic communities are exposed to complex transport and transformation processes. However, detailed process knowledge as revealed by Lagrangian measurements adjusted to travel time is rare in large rivers, in particular at hydrological extremes. To fill this gap, we investigated autotrophic processes, heterotrophic carbon utilization, and micropollutant concentrations applying a Lagrangian sampling design in a 600 km section of the River Elbe (Germany) at historically low discharge. Under base flow conditions, we expect the maximum intensity of instream processes and of point source impacts. Phytoplankton biomass and photosynthesis increased from upstream to downstream sites but maximum chlorophyll concentration was lower than at mean discharge. Concentrations of dissolved macronutrients decreased to almost complete phosphate depletion and low nitrate values. The longitudinal increase of bacterial abundance and production was less pronounced than in wetter years and bacterial community composition changed downstream. Molecular analyses revealed a longitudinal increase of many DOM components due to microbial production, whereas saturated lipid-like DOM, unsaturated aromatics and polyphenols, and some CHOS surfactants declined. In decomposition experiments, DOM components with high O/C ratios and high masses decreased whereas those with low O/C ratios, low masses, and high nitrogen content increased at all sites. Radiocarbon age analyses showed that DOC was relatively old (890-1870 years B.P.), whereas the mineralized fraction was much younger suggesting predominant oxidation of algal lysis products and exudates particularly at downstream sites. Micropollutants determining toxicity for algae (terbuthylazine, terbutryn, isoproturon and lenacil), hexachlorocyclohexanes and DDTs showed higher concentrations from the middle towards the downstream part but calculated toxicity was not negatively correlated to phytoplankton. Overall, autotrophic and heterotrophic process rates and micropollutant concentrations increased from up- to downstream reaches, but their magnitudes were not distinctly different to conditions at medium discharges.

Conditioning Film and Early Biofilm Succession on Plastic Surfaces.

In the context of environmental plastic pollution, it is still under debate if and how the "plastisphere", a plastic-specific microbial community, emerges. In this study, we tested the hypothesis that the first conditioning film of dissolved organic matter (DOM) sorbs selectively to polymer substrates and that microbial attachment is governed in a substrate-dependent manner. We investigated the adsorption of stream water-derived DOM to polyethylene terephthalate (PET), polystyrene (PS), and glass (as control) including UV-weathered surfaces by Fourier-transform ion cyclotron mass spectrometry. Generally, the saturated, high-molecular mass and thus more hydrophobic fraction of the original stream water DOM preferentially adsorbed to the substrates. The UV-weathered polymers adsorbed more polar, hydrophilic OM as compared to the dark controls. The amplicon sequencing data of the initial microbial colonization process revealed a tendency of substrate specificity for biofilm attachment after 24 h and a clear convergence of the communities after 72 h of incubation. Conclusively, the adsorbed OM layer developed depending on the materials' surface properties and increased the water contact angles, indicating higher surface hydrophobicity as compared to pristine surfaces. This study improves our understanding of molecular and biological interactions at the polymer/water interface that are relevant to understand the ecological impact of plastic pollution on a community level.

High concentrations of dissolved biogenic methane associated with cyanobacterial blooms in East African lake surface water.

The contribution of oxic methane production to greenhouse gas emissions from lakes is globally relevant, yet uncertainties remain about the levels up to which methanogenesis can counterbalance methanotrophy by leading to CH4 oversaturation in productive surface waters. Here, we explored the biogeochemical and microbial community variation patterns in a meromictic soda lake, in the East African Rift Valley (Kenya), showing an extraordinarily high concentration of methane in oxic waters (up to 156 µmol L-1). Vertical profiles of dissolved gases and their isotopic signature indicated a biogenic origin of CH4. A bloom of Oxyphotobacteria co-occurred with abundant hydrogenotrophic and acetoclastic methanogens, mostly found within suspended aggregates promoting the interactions between Bacteria, Cyanobacteria, and Archaea. Moreover, aggregate sedimentation appeared critical in connecting the lake compartments through biomass and organic matter transfer. Our findings provide insights into understanding how hydrogeochemical features of a meromictic soda lake, the origin of carbon sources, and the microbial community profiles, could promote methane oversaturation and production up to exceptionally high rates.

Disentangling multiple chemical and non-chemical stressors in a lotic ecosystem using a longitudinal approach.

Meeting ecological and water quality standards in lotic ecosystems is often failed due to multiple stressors. However, disentangling stressor effects and identifying relevant stressor-effect-relationships in complex environmental settings remain major challenges. By combining state-of-the-art methods from ecotoxicology and aquatic ecosystem analysis, we aimed here to disentangle the effects of multiple chemical and non-chemical stressors along a longitudinal land use gradient in a third-order river in Germany. We distinguished and evaluated four dominant stressor categories along this gradient: (1) Hydromorphological alterations: Flow diversity and substrate diversity correlated with the EU-Water Framework Directive based indicators for the quality element macroinvertebrates, which deteriorated at the transition from near-natural reference sites to urban sites. (2) Elevated nutrient levels and eutrophication: Low to moderate nutrient concentrations together with complete canopy cover at the reference sites correlated with low densities of benthic algae (biofilms). We found no more systematic relation of algal density with nutrient concentrations at the downstream sites, suggesting that limiting concentrations are exceeded already at moderate nutrient concentrations and reduced shading by riparian vegetation. (3) Elevated organic matter levels: Wastewater treatment plants (WWTP) and stormwater drainage systems were the primary sources of bioavailable dissolved organic carbon. Consequently, planktonic bacterial production and especially extracellular enzyme activity increased downstream of those effluents showing local peaks. (4) Micropollutants and toxicity-related stress: WWTPs were the predominant source of toxic stress, resulting in a rapid increase of the toxicity for invertebrates and algae with only one order of magnitude below the acute toxic levels. This toxicity correlates negatively with the contribution of invertebrate species being sensitive towards pesticides (SPEARpesticides index), probably contributing to the loss of biodiversity recorded in response to WWTP effluents. Our longitudinal approach highlights the potential of coordinated community efforts in supplementing established monitoring methods to tackle the complex phenomenon of multiple stress.

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.

Short communication on "Differentiating with fluorescence spectroscopy the sources of dissolved organic matter in soils subjected to drying" [Zsolnay et al. Chemosphere 38, 45-50, 1999].

This article is a short, but very important comment to the often used humification index (HIX), assumed to indicate the degree of biochemical degradation of dissolved organic matter in water samples. HIX is commonly calculated by dividing the fluorescence intensity detected at longer wave lengths by the intensity detected at shorter wavelengths. However, we found typos in the original article that affect the used equation directly and thus the results. We compared the different ways to calculate HIX and found a strong correlation between all results. Nevertheless, we recommend to use only equation 1, especially if data from different studies should be compared.

Molecular change of dissolved organic matter and patterns of bacterial activity in a stream along a land-use gradient.

Fluvial networks are globally relevant for the processing of dissolved organic matter (DOM). To investigate the change in molecular DOM diversity along the river course, high-field FTICR mass spectrometry and NMR spectroscopy of riverine DOM as well as bacterial abundance and activity were measured in a third order stream along a land-use gradient from pristine, agricultural to urban landscapes. DOM composition showed a clear evolution along the river course with an initial decrease of average oxidation and unsaturation followed by an increased relative abundance of CHNO and CHOS compounds introduced by agriculture and waste water, respectively. DOM composition was dominated by rather unsaturated CHO compounds (H/C ≤ 1) in headwaters and by more aliphatic molecules at downstream sites. Oxygenated functional groups shifted from aromatic ethers and hydroxyl groups to aliphatic carboxylic acids and aliphatic hydroxyl groups. This massive dislocation of oxygen significantly increased the diversity of atomic environments in branched aliphatic groups from headwater to downstream DOM. Mass spectra of DOM enabled the detection of compositional relationships to bacterial abundance and activity which was positively related to more aliphatic components (H/C > 1) and negatively related to unsaturated components. FTICR mass and NMR spectra corroborated the initial decline in DOM molecular diversity predicted by the River Continuum Concept (RCC) but demonstrated an anthropogenic increase in the molecular diversity of DOM further downstream. While the high DOM molecular diversity in first order headwater streams was the result of small scale ecosystem plurality, agriculture and waste water treatment introduced many components in the lower reaches. These anthropogenic influences together with massive bacterial oxidation of DOM contributed to a growth of molecular diversity of downstream DOM whose composition and structure differed entirely from those found in pristine headwaters.

Metallophore profiling of nitrogen-fixing Frankia spp. to understand metal management in the rhizosphere of actinorhizal plants.

Frankia spp. are widespread nitrogen-fixing soil bacteria, which often live in symbiosis with a broad range of hosts. Metal homeostasis plays a crucial role in the success of the symbiosis regarding the acquisition of essential trace metals and detoxification of potentially toxic elements. We have hypothesised that Frankia releases many organic ligands with a broad spectrum of affinity for essential and toxic metals. We coined the term 'ligandosphere' to describe the entirety of excreted metal complexing agents and ligands derived from the dissolved organic matter. Using metal isotope-coded profiling (MICP); metallophores of physiological important and toxic trace metals were identified by the addition of stable metal isotope pairs such as 54Fe/58Fe, 63Cu/65Cu, 66Zn/68Zn or 95Mo/98Mo. Liquid chromatography coupled to a mass spectrometer revealed strong variations of the metallophore profile in between the 14 test-strains. In total, about 83 organic ligands were identified as binding to one of the tested metals. The predicted sum formula of the major Fe binding ligands and MS/MS experiments suggested that several metallophore candidates have a similar molecular backbone. Growth experiments with a hyper-producer of metallophores revealed a positive relationship between metallophore production and the concentration of Cu in the growth medium. The present study provides the first comprehensive overview of the complexity of Frankia's ligandosphere. It opens a path to a deeper understanding of mechanisms that regulate metal homeostasis in frankiae. Deciphering these mechanisms is important since the fitness of actinorhizal plants and their potential in ecological restoration relies heavily on their symbiosis with frankiae.

Redox Conditions Affect Dissolved Organic Carbon Quality in Stratified Freshwaters.

The quality of dissolved organic carbon (DOC) affects both carbon cycling in surface waters and drinking water production. Not much is known about the influence of environmental conditions on DOC quality. We studied the effect of redox conditions on the chemical composition of DOC in a drinking water reservoir by Fourier transform ion cyclotron resonance mass spectrometry in combination with sediment core incubation experiments under manipulated redox conditions. We observed clear differences in DOC quality among oxic epilimnion, anoxic hypolimnion, and sediment porewater. Sediment porewater showed relatively high intensities of polyphenol-like components with H/C ratios of <1 and O/C ratios of >0.6. Consistent with this, anoxic incubation of a sediment core resulted in an accumulation of these components in the overlying water. The observed pattern of DOC quality change can be explained by redox-dependent adsorption/desorption of DOC on iron minerals. Under oxic conditions, the polyphenol-like components bind on freshly formed iron hydroxides, a process that affects both DOC stability in surface waters and treatability during drinking water production.

A new approach for evaluating transformations of dissolved organic matter (DOM) via high-resolution mass spectrometry and relating it to bacterial activity.

Streams are important sites of transformation of dissolved organic matter (DOM). The molecular characterization of DOM-quality changes requires sophisticated analytical evaluation techniques. The goal of our study was to link molecular DOM transformation with bacterial activity. We measured the degradation of leaf leachate over a gradient of bacterial production obtained by different rates of percolation of sediments in seven experimental flumes on five sampling dates. We developed a new strategy for evaluating molecular formula data sets obtained by ultra-high resolution Fourier transform ion cyclotron resonance mass spectrometry (FTICR MS), in which the time-dependent change of component abundance was fitted by a linear regression model after normalization of mass peak intensities. All components were categorized by calculating the slope (change of percent intensity per day) in each of the seven flumes. These slopes were then related to cumulative bacterial production. The concentration of DOM decreased quickly in all flumes. Bacterial activity was higher in flumes with percolated sediment than in those without percolation, whereas plankton bacterial activity was higher in flumes without percolation or without sediment. There were no differences in molecular-DOM characteristics between flumes, but there were distinct changes over time. Positive slopes, i.e. increasing intensities over time, were found for small molecules (MW < 450 Da) and high O/C ratios, whereas decreasing intensities were observed less often and only for large molecules and low O/C ratios. The positive slopes of produced components showed a positive relationship to bacterial production for small and for oxygen-rich components. The negative slopes of degraded components were negatively related to bacterial production for large and for oxygen-deficient molecules. Overall, the approach provided new insights into the transformation of specific molecular DOM components.

Land-based salmon aquacultures change the quality and bacterial degradation of riverine dissolved organic matter.

Aquacultures are of great economic importance worldwide but pollute pristine headwater streams, lakes, and estuaries. However, there are no in-depth studies of the consequences of aquacultures on dissolved organic matter (DOM) composition and structure. We performed a detailed molecular level characterization of aquaculture DOM quality and its bacterial degradation using four salmon aquacultures in Chile. Fluorescence measurements, ultrahigh-resolution mass spectrometry, and nuclear magnetic resonance spectroscopy of the DOM revealed specific and extensive molecular alterations caused by aquacultures. Aquacultures released large quantities of readily bioavailable metabolites (primarily carbohydrates and peptides/proteins, and lipids), causing the organic matter downstream of all the investigated aquacultures to deviate strongly from the highly processed, polydisperse and molecularly heterogeneous DOM found in pristine rivers. However, the upstream individual catchment DOM signatures remained distinguishable at the downstream sites. The benthic algal biovolume decreased and the bacterial biovolume and production increased downstream of the aquacultures, shifting stream ecosystems to a more heterotrophic state and thus impairing the ecosystem health. The bacterial DOM degradation rates explain the attenuation of aquaculture DOM within the subsequent stream reaches. This knowledge may aid the development of improved waste processing facilities and may help to define emission thresholds to protect sensitive stream ecosystems.

Differences in DOM of rewetted and natural peatlands - Results from high-field FT-ICR-MS and bulk optical parameters.

Peatlands can be a potential source of dissolved organic matter (DOM) in fresh water catchment areas. The quantity and quality of DOM can differ between pristine, degraded and rewetted peatlands. Due to the large scale and continuing losses of peatlands, their conservation and restoration has been increasingly emphasized. Mostly rewetting measures are required to improve the hydrology of damaged peatlands, which is a precondition for the resettlement of peat-forming plant species. Thus, in term of DOM, there is a special need to understand how rewetting measures affect DOM characteristics and concentrations. To estimate the potential leaching of humic substances from rewetted areas two natural sites were compared with four artificially rewetted sites in a peatland area of the Harz Mountains National Park, Germany. This was done with regards to DOM quality by combining the results from Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS, measured at one time in Spring) and excitation-emission-matrix fluorescence spectroscopy (EEMF, measured monthly for the period of one year). The DOM quality was significantly less variable in the pristine peatland soil water compared to the rewetted peatland soil waters, from both a spatial and a seasonal perspective. The soil water from the rewetted peatland sites showed a higher degree of humification compared to pristine peatland. DOC concentration was mostly consistent in the pristine peatland over the year. The rewetted peatlands showed higher DOC levels in Summer months and lower DOC in Winter months compared to the pristine peatland. It can be concluded that the rewetting of peatlands is coupled with high concentrations of DOC in soil water and its quality is highly aromatic (as reflected by the observed values from the humification index) during times with elevated temperature. The results may have a significant input for dynamic catchment area studies with regards to rewetting peatland sites.

Discharge determines production of, decomposition of and quality changes in dissolved organic carbon in pre-dams of drinking water reservoirs.

Pre-dams are small reservoirs constructed upstream of the main drinking water reservoirs and are used for nutrient removal and sediment trapping. Little is known about the role of pre-dams regarding the production and decomposition of dissolved organic carbon (DOC) in relation to discharge and how this affects the quality of DOC in the water. We combined quantitative and qualitative investigations under different hydrological conditions at three pre-dams exhibiting a gradient from oligotrophic/high-DOC to eutrophic/low-DOC. All pre-dams were mainly autotrophic in their upper water layers. The ratio of OC production to total gained OC (i.e. OC import+OC production) decreased with increasing discharge. On average, 0-30% of the total gained OC was produced within the pre-dams. The amount of microbially decomposed DOC increased with the average water residence time (WRT) and with the trophic status of the pre-dams. Radiocarbon analyses of respired CO2 revealed that heterotrophic bacteria preferentially utilized old DOC components (195-395years before present) under base flow conditions, whereas younger components (modern, i.e. OC produced after 1950) were utilized at high discharge. DOC quality changed significantly over the year within the pre-dams: High proportions of algae-derived DOC were observed during base flow in summer, and the freshness index (ß/α ratio) decreased significantly with higher discharges. DOC production and quality changes in response to hydrological conditions should be considered for future water quality management in reservoirs, as climate scenarios for temperate regions predict decreased runoffs leading to longer WRT and increased eutrophication and production of algae-derived OC.

Selectivity of solid phase extraction of freshwater dissolved organic matter and its effect on ultrahigh resolution mass spectra.

Solid phase extraction (SPE) is often used for enrichment and clean-up prior to analysis of dissolved organic matter (DOM) by electrospray ionization (ESI) coupled to ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS). It is generally accepted that extraction by SPE is not quantitative with respect to carbon concentration. However, little information is available on the selectivity of different SPE sorbents and the resulting effect for the acquired DOM mass spectra. Freshwater samples were extracted by the widely used PPL, HLB and C18 sorbents and the molecular composition and size distribution of the DOM in the extracts and in the permeates was compared to the original sample. Dissolved organic carbon (DOC) recoveries ranged between 20% and 65% for the three tested SPE sorbents. Size-exclusion chromatography coupled to organic carbon detection (SEC-OCD) revealed that limited recovery by PPL and HLB was primarily due to incomplete elution of a fraction of apparent high molecular weight from the solid phase. In contrast, incomplete retention on the solid phase, mainly observed for the C18 cartridge, was attributed to a fraction of low molecular weight. The FT-ICR mass spectra of the original sample and the SPE extracts did not differ significantly in their molecular weight distribution, but they showed sorbent specific differences in the degree of oxygenation and saturation. We concluded that the selective enrichment of freshwater DOM by SPE is less critical for subsequent FT-ICR MS analysis, because those fractions that are not sufficiently recovered have comparatively small effects on the mass spectra. This was confirmed by the extraction of model compounds, showing that very polar and small molecules are poorly extracted, but also have a low response in ESI-MS. Of the three tested SPE cartridges the PPL material offered the best properties for DOM enrichment for subsequent FT-ICR MS analysis as it minimizes too strong and too weak DOM-sorbent interactions.

Molecular formula assignment for dissolved organic matter (DOM) using high-field FT-ICR-MS: chemical perspective and validation of sulphur-rich organic components (CHOS) in pit lake samples.

Molecular formula assignment is one of the key challenges in processing high-field Fourier transform ion cyclotron resonance mass spectrometric (FT-ICR-MS) datasets. The number of potential solutions for an elemental formula increases exponentially with increasing molecular mass, especially when non-oxygen heteroatoms like N, S or P are included. A method was developed from the chemical perspective and validated using a Suwannee River Fulvic Acid (SRFA) dataset which is dominated by components consisting exclusively of C, H and O (78 % CHO). In order to get information on the application range and robustness of this method, we investigated a FT-ICR-MS dataset which was merged from 18 mine pit lake pore waters and 3 river floodplain soil waters. This dataset contained 50 % CHO and 18 % CHOS on average, whereas the former SRFA dataset contained only 1.5 % CHOS. The mass calculator was configured to allow up to five nitrogen atoms and up to one sulphur atom in assigning formulas to mass peaks. More than 50 % multiple-formula assignments were found for peaks with masses > 650 Da. Based on DBE - O frequency diagrams, many CHO, CHOS1, CHON1 and CHON1S1 molecular series were ultimately assigned to many m/z and considered to be reliable solutions. The unequivocal data pool could thus be enlarged by 523 (6.8 %) CHOS1 components. In contrast to the method validation with CHO-rich SRFA, validation with sulphur-rich pit lake samples showed that formulas with a higher number of non-oxygen heteroatoms can be more reliable assignments in many cases. As an example: CHOS molecular series were reliable and the CHO classes were unreliable amongst other molecular classes in many multiple-formula assignments from the sulphur-rich pit lake samples. Graphical abstract An exemplary frequency versus DBE - O diagram. CHOS components but not CHO (and not CHON2 or CHON2S) components were considered here reliable.

Bacterial production and their role in the removal of dissolved organic matter from tributaries of drinking water reservoirs.

Enhanced concentrations of dissolved organic matter (DOM) in freshwaters are an increasing problem in drinking water reservoirs. In this study we investigated bacterial DOM degradation rates in the tributaries of the reservoirs and tested the hypotheses that (1) DOM degradation is high enough to decrease DOM loads to reservoirs considerably, (2) DOM degradation is affected by stream hydrology, and (3) phosphorus addition may stimulate bacterial DOM degradation. Bacterial biomass production, which was used as a measure of DOM degradation, was highest in summer, and was usually lower at upstream than at downstream sites. An important proportion of bacterial production was realized in epilithic biofilms. Production of planktonic and biofilm bacteria was related to water temperature. Planktonic production weakly correlated to DOM quality and to total phosphorus concentration. Addition of soluble reactive phosphorus did not stimulate bacterial DOM degradation. Overall, DOM was considerably degraded in summer at low discharge levels, whereas degradation was negligible during flood events (when DOM load in reservoirs was high). The ratio of DOM degradation to total DOM release was negatively related to discharge. On annual average, only 0.6-12% of total DOM released by the catchments was degraded within the tributaries.

High-field FTICR-MS data evaluation of natural organic matter: are CHON5S2 molecular class formulas assigned to (13)C isotopic m/z and in reality CHO components?

The analysis of dissolved organic matter (DOM) using high-field Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) poses challenges in molecular formula assignment. The consideration of (13)C isotopes provides new insights into the consistent elemental formula solutions. Modern software helps to overcome misinterpretation, but false assignments of molecular classes to mass peaks have rarely been elucidated until now. It will be demonstrated that this can be important with formula assignments comprising exactly five nitrogen and two sulfur atoms in DOM data sets: the molecular class CHON5S2. The existence of such components in DOM under tripeptide Met-His-Cys formed with the formula C14H23O4N5S2 cannot be excluded; however, components containing 5 N and 2 S should be suspected to not be highly abundant. The true elemental compositions of such unusual "N5S2 moieties" were calculated using Suwannee River fulvic acid (SRFA) data from the literature and one data set from acidic pit lake pore water. The replacement of a H3N5S2 moiety with a (13)C1(12)C5O4 moiety explained more than 95% of the questionable "N5S2 moieties". This finding was proved by calculation of δ(13)C‰ values from relative peak intensities.